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Sample records for preischaemic erythropoietin administration

  1. Erythropoietin.

    PubMed

    Bunn, H Franklin

    2013-03-01

    During the past century, few proteins have matched erythropoietin (Epo) in capturing the imagination of physiologists, molecular biologists, and, more recently, physicians and patients. Its appeal rests on its commanding role as the premier erythroid cytokine, the elegant mechanism underlying the regulation of its gene, and its remarkable impact as a therapeutic agent, arguably the most successful drug spawned by the revolution in recombinant DNA technology. This concise review will begin with a synopsis of the colorful history of this protein, culminating in its purification and molecular cloning. It then covers in more detail the contemporary understanding of Epo's physiology as well as its structure and interaction with its receptor. A major part of this article focuses on the regulation of the Epo gene and the discovery of HIF, a transcription factor that plays a cardinal role in molecular adaptation to hypoxia. In the concluding section, a synopsis of Epo's role in disorders of red blood cell production will be followed by an assessment of the remarkable impact of Epo therapy in the treatment of anemias, as well as concerns that provide a strong impetus for the development of even safer and more effective treatment. PMID:23457296

  2. Erythropoietin.

    PubMed

    Jelkmann, Wolfgang

    2016-01-01

    Total hemoglobin (Hb) mass is an important determinant of aerobic power. The glycoprotein erythropoietin (Epo) promotes the production of red blood cells (RBCs). The present article reviews the regulation of erythropoiesis and ways of its manipulation. The various Epos, e.g. recombinant human (rh)Epo and (epoetin), and their long-acting analogues can be misused by cheating athletes, but the drugs are detectable by chemical tests, because their glycan isoform structures differ from those of endogenous Epo. Still, anti-doping control has become more difficult, since additional erythropoiesis-stimulating agents have become available (Epo mimetics, activin inhibitors, and small-molecule chemical drugs activating EPO expression). A major problem is created by hypoxia-inducible factor (HIF) stabilizers (e.g. α-ketoglutarate competitors and Co2+ salt) which activate HIFs and thus increase EPO expression. Direct EPO transfer is theoretically also possible but medically little advanced. To overcome weaknesses of direct testing of biological fluids, the World Anti-Doping Agency has implemented the Athlete Biological Passport for continuous monitoring of RBC parameters of athletes. Blood doping is assumed when distinct parameters (blood Hb concentration and reticulocytes) change in a nonphysiological way. PMID:27348128

  3. Effects of Erythropoietin Administration on Adrenal Glands of Landrace/Large White Pigs after Ventricular Fibrillation.

    PubMed

    Faa, Armando; Faa, Gavino; Papalois, Apostolos; Obinu, Eleonora; Locci, Giorgia; Pais, Maria Elena; Lelovas, Pavlos; Barouxis, Dimitrios; Pantazopoulos, Charalampos; Vasileiou, Panagiotis V; Iacovidou, Nicoletta; Xanthos, Theodoros

    2016-01-01

    Aim. To evaluate the effects of erythropoietin administration on the adrenal glands in a swine model of ventricular fibrillation and resuscitation. Methods. Ventricular fibrillation was induced via pacing wire forwarded into the right ventricle in 20 female Landrace/Large White pigs, allocated into 2 groups: experimental group treated with bolus dose of erythropoietin (EPO) and control group which received normal saline. Cardiopulmonary resuscitation (CPR) was performed immediately after drug administration as per the 2010 European Resuscitation Council (ERC) guidelines for Advanced Life Support (ALS) until return of spontaneous circulation (ROSC) or death. Animals who achieved ROSC were monitored, mechanically ventilated, extubated, observed, and euthanized. At necroscopy, adrenal glands samples were formalin-fixed, paraffin-embedded, and routinely processed. Sections were stained with hematoxylin-eosin. Results. Oedema and apoptosis were the most frequent histological changes and were detected in all animals in the adrenal cortex and in the medulla. Mild and focal endothelial lesions were also detected. A marked interindividual variability in the degree of the intensity of apoptosis and oedema at cortical and medullary level was observed within groups. Comparing the two groups, higher levels of pathological changes were detected in the control group. No significant difference between the two groups was observed regarding the endothelial changes. Conclusions. In animals exposed to ventricular fibrillation, EPO treatment has protective effects on the adrenal gland. PMID:27504455

  4. Effects of Erythropoietin Administration on Adrenal Glands of Landrace/Large White Pigs after Ventricular Fibrillation

    PubMed Central

    Faa, Gavino; Papalois, Apostolos; Obinu, Eleonora; Locci, Giorgia; Pais, Maria Elena; Lelovas, Pavlos; Barouxis, Dimitrios; Pantazopoulos, Charalampos; Vasileiou, Panagiotis V.; Iacovidou, Nicoletta; Xanthos, Theodoros

    2016-01-01

    Aim. To evaluate the effects of erythropoietin administration on the adrenal glands in a swine model of ventricular fibrillation and resuscitation. Methods. Ventricular fibrillation was induced via pacing wire forwarded into the right ventricle in 20 female Landrace/Large White pigs, allocated into 2 groups: experimental group treated with bolus dose of erythropoietin (EPO) and control group which received normal saline. Cardiopulmonary resuscitation (CPR) was performed immediately after drug administration as per the 2010 European Resuscitation Council (ERC) guidelines for Advanced Life Support (ALS) until return of spontaneous circulation (ROSC) or death. Animals who achieved ROSC were monitored, mechanically ventilated, extubated, observed, and euthanized. At necroscopy, adrenal glands samples were formalin-fixed, paraffin-embedded, and routinely processed. Sections were stained with hematoxylin-eosin. Results. Oedema and apoptosis were the most frequent histological changes and were detected in all animals in the adrenal cortex and in the medulla. Mild and focal endothelial lesions were also detected. A marked interindividual variability in the degree of the intensity of apoptosis and oedema at cortical and medullary level was observed within groups. Comparing the two groups, higher levels of pathological changes were detected in the control group. No significant difference between the two groups was observed regarding the endothelial changes. Conclusions. In animals exposed to ventricular fibrillation, EPO treatment has protective effects on the adrenal gland. PMID:27504455

  5. Effects of Erythropoietin Administration on Adrenal Glands of Landrace/Large White Pigs after Ventricular Fibrillation.

    PubMed

    Faa, Armando; Faa, Gavino; Papalois, Apostolos; Obinu, Eleonora; Locci, Giorgia; Pais, Maria Elena; Lelovas, Pavlos; Barouxis, Dimitrios; Pantazopoulos, Charalampos; Vasileiou, Panagiotis V; Iacovidou, Nicoletta; Xanthos, Theodoros

    2016-01-01

    Aim. To evaluate the effects of erythropoietin administration on the adrenal glands in a swine model of ventricular fibrillation and resuscitation. Methods. Ventricular fibrillation was induced via pacing wire forwarded into the right ventricle in 20 female Landrace/Large White pigs, allocated into 2 groups: experimental group treated with bolus dose of erythropoietin (EPO) and control group which received normal saline. Cardiopulmonary resuscitation (CPR) was performed immediately after drug administration as per the 2010 European Resuscitation Council (ERC) guidelines for Advanced Life Support (ALS) until return of spontaneous circulation (ROSC) or death. Animals who achieved ROSC were monitored, mechanically ventilated, extubated, observed, and euthanized. At necroscopy, adrenal glands samples were formalin-fixed, paraffin-embedded, and routinely processed. Sections were stained with hematoxylin-eosin. Results. Oedema and apoptosis were the most frequent histological changes and were detected in all animals in the adrenal cortex and in the medulla. Mild and focal endothelial lesions were also detected. A marked interindividual variability in the degree of the intensity of apoptosis and oedema at cortical and medullary level was observed within groups. Comparing the two groups, higher levels of pathological changes were detected in the control group. No significant difference between the two groups was observed regarding the endothelial changes. Conclusions. In animals exposed to ventricular fibrillation, EPO treatment has protective effects on the adrenal gland.

  6. Erythropoietin administration partially prevents adipose tissue loss in experimental cancer cachexia models

    PubMed Central

    Penna, Fabio; Busquets, Silvia; Toledo, Miriam; Pin, Fabrizio; Massa, David; López-Soriano, Francisco J.; Costelli, Paola; Argilés, Josep M.

    2013-01-01

    Cancer-associated cachexia is characterized, among other symptoms, by a dramatic loss of both muscle and fat. In addition, the cachectic syndrome is often associated with anemia. The object of the present investigation was to assess the effects of erythropoietin (EPO) treatment on experimental cancer cachexia models. The results clearly show that, in addition to the improvement of the hematocrit, EPO treatment promoted a partial preservation of adipose tissue while exerting negligible effects on muscle loss. Administration of EPO to tumor-bearing animals resulted in a significant increase of lipoprotein lipase (LPL) activity in adipose tissue, suggesting that the treatment favored triacylglycerol (TAG) accumulation in the adipose tissue. In vitro experiments using both adipose tissue slices and 3T3-L1 adipocytes suggests that EPO is able to increase the lipogenic rate through the activation of its specific receptor (EPOR). This metabolic pathway, in addition to TAG uptake by LPL, may contribute to the beneficial effects of EPO on fat preservation in cancer cachexia. PMID:23966665

  7. A biomathematical model of human erythropoiesis under erythropoietin and chemotherapy administration.

    PubMed

    Schirm, Sibylle; Engel, Christoph; Loeffler, Markus; Scholz, Markus

    2013-01-01

    Anaemia is a common haematologic side effect of dose-dense multi-cycle cytotoxic polychemotherapy requiring erythrocyte transfusions or erythropoietin (EPO) administration. To simulate the effectiveness of different EPO application schedules, we performed both modelling of erythropoiesis under chemotherapy and pharmacokinetic and dynamic modelling of EPO applications in the framework of a single comprehensive biomathematical model. For this purpose, a cell kinetic model of bone marrow erythropoiesis was developed that is based on a set of differential compartment equations describing proliferation and maturation of erythropoietic cell stages. The system is regulated by several feedback loops comprising those mediated by EPO. We added a model of EPO absorption after injection at different sites and a pharmacokinetic model of EPO derivatives to account for the effects of external EPO applications. Chemotherapy is modelled by a transient depletion of bone marrow cell stages. Unknown model parameters were determined by fitting the predictions of the model to data sets of circulating erythrocytes, haemoglobin, haematocrit, percentage of reticulocytes or EPO serum concentrations derived from the literature or cooperating clinical study groups. Parameter fittings resulted in a good agreement of model and data. Depending on site of injection and derivative (Alfa, Beta, Delta, Darbepoetin), nine groups of EPO applications were distinguished differing in either absorption kinetics or pharmacokinetics. Finally, eight different chemotherapy protocols were modelled. The model was validated on the basis of scenarios not used for parameter fitting. Simulations were performed to analyze the impact of EPO applications on the risk of anaemia during chemotherapy. We conclude that we established a model of erythropoiesis under chemotherapy that explains a large set of time series data under EPO and chemotherapy applications. It allows predictions regarding yet untested EPO schedules

  8. Differential effects of erythropoietin on neural and cognitive measures of executive function 3 and 7 days post-administration.

    PubMed

    Miskowiak, Kamilla; Inkster, Becky; O'Sullivan, Ursula; Selvaraj, Sudhakar; Goodwin, Guy M; Harmer, Catherine J

    2008-01-01

    Erythropoietin (Epo) has neuroprotective and neurotrophic effects and improves cognitive function in animal models of neurodegenerative and neuropsychiatric illness. In humans, weekly Epo administration over 3 months improves cognitive function in schizophrenia. The neural underpinnings and time-course of this effect of Epo are currently unknown. It is also unclear whether the cognitive improvement reflects direct neurobiological actions or is secondary to hematological effects. We therefore assessed the actions of single administration of Epo (40,000 IU) vs. saline to healthy volunteers on cognitive and neural measures of executive function using a verbal fluency task and N-back working memory (WM) paradigm during functional magnetic resonance imaging (fMRI) on day 3 and 7 after administration in two separate cohorts of subjects. Epo modulated neuronal response in a fronto-parietal network during WM performance at both time points; on day 3 after administration, activation was increased in left-hemisphere frontal and cingulate cortex and reduced in the right parietal cortex; in contrast, neural response was enhanced in a right-lateralized fronto-parietal network and reduced in left-side regions 1 week post-administration. In addition, Epo-treated volunteers displayed improved verbal fluency performance 1 week post-administration. These effects occurred in the absence of changes in hematological measures suggesting that they reflect direct neurobiological actions of Epo. The findings are consistent with enduring effects of Epo on neurotrophic signaling and induction of neurochemical changes over time in neural networks typically affected in neuropsychiatric illness. The present study supports the notion that Epo may have clinical applications in the treatment of psychiatric disorder characterized by cognitive dysfunction.

  9. A mouse model of adult-onset anaemia due to erythropoietin deficiency.

    PubMed

    Yamazaki, Shun; Souma, Tomokazu; Hirano, Ikuo; Pan, Xiaoqing; Minegishi, Naoko; Suzuki, Norio; Yamamoto, Masayuki

    2013-01-01

    Erythropoietin regulates erythropoiesis in a hypoxia-inducible manner. Here we generate inherited super-anaemic mice (ISAM) as a mouse model of adult-onset anaemia caused by erythropoietin deficiency. ISAM express erythropoietin in the liver but lack erythropoietin production in the kidney. Around weaning age, when the major erythropoietin-producing organ switches from the liver to the kidney, ISAM develop anaemia due to erythropoietin deficiency, which is curable by administration of recombinant erythropoietin. In ISAM severe chronic anaemia enhances transgenic green fluorescent protein and Cre expression driven by the complete erythropoietin-gene regulatory regions, which facilitates efficient labelling of renal erythropoietin-producing cells. We show that the majority of cortical and outer medullary fibroblasts have the innate potential to produce erythropoietin, and also reveal a new set of erythropoietin target genes. ISAM are a useful tool for the evaluation of erythropoiesis-stimulating agents and to trace the dynamics of erythropoietin-producing cells. PMID:23727690

  10. Erythropoietin Derived by Chemical Synthesis

    PubMed Central

    Shieh, Jae-Hung; Peguero, Elizabeth; Hendrickson, Ronald; Moore, Malcolm A. S.; Danishefsky, Samuel J.

    2014-01-01

    Erythropoietin is a signaling glycoprotein that controls the fundamental process of erythropoiesis, orchestrating the production and maintenance of red blood cells. As administrated clinically, erythropoietin has a polypeptide backbone with complex dishomogeneity in its carbohydrate domains. Here we describe the total synthesis of homogeneous erythropoietin with consensus carbohydrate domains incorporated at all of the native glycosylation sites. The oligosaccharide sectors were built by total synthesis and attached stereospecifically to peptidyl fragments of the wild-type primary sequence, themselves obtained by solid-phase peptide synthesis. The glycopeptidyl constructs were joined by chemical ligation, followed by metal-free dethiylation, and subsequently folded. This homogeneous erythropoietin glycosylated at the three wild-type aspartates with N-linked high-mannose sialic acid–containing oligosaccharides and O-linked glycophorin exhibits Procrit-level in vivo activity in mice. PMID:24337294

  11. Activity increase in EpoR and Epo expression by intranasal recombinant human erythropoietin (rhEpo) administration in ischemic hippocampi of adult rats.

    PubMed

    Castañeda-Arellano, R; Feria-Velasco, A I; Rivera-Cervantes, M C

    2014-11-01

    Erythropoietin in the nervous system is a potential neuroprotective factor for cerebral ischemic damage due to specific-binding to the erythropoietin receptor, which is associated with survival mechanisms. However, the role of its receptor is unclear. Thus, this work assessed whether a low dose (500UI/Kg) of intranasal recombinant human erythropoietin administered 3h after ischemia induced changes in the activation of its receptor at the Tyr456-phosphorylated site in ischemic hippocampi in rats. The results showed that recombinant human erythropoietin after injury maintained cell survival and was associated with an increase in receptor phosphorylation at the Tyr456 site as an initial signaling step, which correlated with a neuroprotective effect. PMID:25219375

  12. The Impact of Tumor Expression of Erythropoietin Receptors and Erythropoietin on Clinical Outcome of Esophageal Cancer Patients Treated With Chemoradiation

    SciTech Connect

    Rades, Dirk Golke, Helmut; Schild, Steven E.; Kilic, Ergin

    2008-05-01

    Background: To investigate the impact of tumor erythropoietin receptors (Epo-R) and erythropoietin (Epo) expression in 64 patients with Stage III esophageal cancer receiving or not receiving erythropoietin during chemoradiation. Materials and Methods: The impact of tumor Epo-R expression, Epo expression, and 10 additional factors (age, Karnofsky-Performance-Score [KPS], tumor length, T and N stage, histology and grading, hemoglobin during radiotherapy, erythropoietin administration, surgery) on overall survival (OS) and locoregional control (LC) was evaluated. Results: Improved OS was associated with low ({<=}20%) Epo expression (p = 0.049), KPS >80 (p 0.008), T3 stage (p = 0.010), hemoglobin {>=}12 g/dL (p < 0.001), and surgery (p = 0.010). Erythropoietin receptor expression showed a trend (p = 0.09). Locoregional control was associated with T stage (p = 0.005) and hemoglobin (p < 0.001), almost with erythropoietin administration (p = 0.06). On multivariate analyses, OS was associated with KPS (p = 0.045) and hemoglobin (p = 0.032), LC with hemoglobin (p < 0.001). Patients having low expression of both Epo-R and Epo had better OS (p = 0.003) and LC (p = 0.043) than others. Two-year OS was nonsignificantly better (p = 0.25) in patients with low Epo-R expression receiving erythropoietin (50%) than in those with higher Epo-R expression receiving erythropoietin (21%), low Epo-R expression/no erythropoietin administration (29%), or higher Epo-R expression/no erythropoietin administration (18%). Two-year LC rates were, respectively, 65%, 31%, 26%, and 29% (p = 0.20). Results for Epo expression were similar. Conclusions: Higher Epo-R expression or Epo expression seemed to be associated with poorer outcomes. Patients with low expression levels receiving erythropoietin seemed to do better than patients with higher expression levels or not receiving erythropoietin. The data need to be confirmed in a larger series of patients.

  13. Erythropoietin and Neonatal Neuroprotection.

    PubMed

    Juul, Sandra E; Pet, Gillian C

    2015-09-01

    Certain groups of neonates are at high risk of developing long-term neurodevelopmental impairment and might be considered candidates for neuroprotective interventions. This article explores some of these high-risk groups, relevant mechanisms of brain injury, and specific mechanisms of cellular injury and death. The potential of erythropoietin (Epo) to act as a neuroprotective agent for neonatal brain injury is discussed. Clinical trials of Epo neuroprotection in preterm and term infants are updated. PMID:26250911

  14. Erythropoietin binding protein from mammalian serum

    DOEpatents

    Clemons, G.K.

    1997-04-29

    Purified mammalian erythropoietin binding-protein is disclosed, and its isolation, identification, characterization, purification, and immunoassay are described. The erythropoietin binding protein can be used for regulation of erythropoiesis by regulating levels and half-life of erythropoietin. A diagnostic kit for determination of level of erythropoietin binding protein is also described. 11 figs.

  15. Erythropoietin binding protein from mammalian serum

    DOEpatents

    Clemons, Gisela K.

    1997-01-01

    Purified mammalian erythropoietin binding-protein is disclosed, and its isolation, identification, characterization, purification, and immunoassay are described. The erythropoietin binding protein can be used for regulation of erythropoiesis by regulating levels and half-life of erythropoietin. A diagnostic kit for determination of level of erythropoietin binding protein is also described.

  16. Renal Cell Protection of Erythropoietin beyond Correcting The Anemia in Chronic Kidney Disease Patients.

    PubMed

    Nasri, Hamid

    2014-01-01

    -erythropoietic tissue/ organ protective efficacy of erythropoietin has become evident, especially in the kidneys (5-12). Various investigations have shown the kidney protective property of erythropoietin in acute kidney injury. In a study to evaluate the ameliorative effects of erythropoietin on renal tubular cells, we studied 40 male rats. We found that erythropoietin was able to prevent the increase in serum creatinine and blood urea nitrogen. Furthermore, co-administration of gentamicin and erythropoietin effectively reduced kidney tissue damage compared to the control group. However, the protective properties of erythropoietin were also evident in our study. When the drug was applied after gentamicin- induced tubular damage we were able to show that the drug was still effective after tissue injury onset. This indicates that erythropoietin may have curative effects in addition to its preventive properties (13). Thus, erythropoietin is a promising kidney protective agent to prevent, ameliorate or attenuate tubular damage induced by gentamicin or other nephrotoxic agents that act in a similar manner to this drug (14-17). Recent studies have elucidated the cellular mechanism involved in kidney erythropoietin production and the consequent events that lead to kidney fibrosis, showing that they are closely related to each other (18-20). In contrast to previous findings, fibroblasts originating from damaged renal tubular epithelial cells do not have an important role in kidney fibrosis, but renal erythropoietin- producing cells, stemming from neural crests, have been shown to trans-differentiate into myofibroblasts after long-term exposure to inflammatory situations related to kidney fibrosis. In fact, almost all myofibroblasts expressing α-smooth muscle actin originate from renal erythropoietin-producing cells, which are naturally peritubular interstitial fibroblastic cells expressing neural cell marker genes but not α-smooth muscle actin. Macrophages and myofibroblasts are responsible

  17. Human cytomegalovirus inhibits erythropoietin production.

    PubMed

    Butler, Lynn M; Dzabic, Mensur; Bakker, Frank; Davoudi, Belghis; Jeffery, Hannah; Religa, Piotr; Bojakowski, Krzysztof; Yaiw, Koon-Chu; Rahbar, Afsar; Söderberg-Naucler, Cecilia

    2014-08-01

    Anemia is a feature of CKD and a complication of renal transplantation, often caused by impaired production of erythropoietin. The kidney is a target organ for human cytomegalovirus (hCMV) in such patients, but it is not known whether hCMV effects erythropoietin production. We found that kidneys from patients with CKD were positive for hCMV protein and that blood levels of hCMV IgG inversely correlated with red blood cell count. In mice, systemic murine cytomegalovirus infection decreased serum erythropoietin levels. In human erythropoietin-producing cells, hCMV inhibited hypoxia-induced expression of erythropoietin mRNA and protein. hCMV early gene expression was responsible, as ultraviolet-inactivated virus had no effect and valganciclovir treatment showed that late gene expression was nonessential. Hypoxia-induced gene transcription is controlled by the transcription factors hypoxia-inducible transcription factor (HIF)-1α and HIF2α, which are constitutively produced but stable only under low oxygen conditions. We found that hCMV inhibited constitutive production of HIF2α mRNA. HIF2α is thought to be the master regulator of erythropoietin transcription. Single-cell analysis revealed that nuclear accumulation of HIF2α was inhibited in hCMV-infected cells, and the extent of inhibition correlated with hCMV protein expression. Our findings suggest that renal hCMV infection could induce or exacerbate anemia in patients.

  18. Recombinant Human Erythropoietin

    PubMed Central

    Bartels, Claudia; Späte, Kira; Krampe, Henning

    2008-01-01

    Treatment of multiple sclerosis (MS) is still unsatisfactory and essentially non-existing for the progressive course of the disease. Recombinant human erythropoietin (EPO) may be a promising neuroprotective/neuroregenerative treatment of MS. In the nervous system, EPO acts anti-apoptotic, antioxidative, anti-inflammatory, neurotrophic and plasticity-modulating. Beneficial effects have been shown in animal models of various neurological and psychiatric diseases, including different models of experimental autoimmune encephalomyelitis. EPO is also effective in human brain disease, as shown in double-blind placebo-controlled clinical studies on ischemic stroke and chronic schizophrenia. An exploratory study on chronic progressive MS yielded lasting improvement in motor and cognitive performance upon high-dose long-term EPO treatment. PMID:21180577

  19. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  20. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  1. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  2. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  3. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  4. Red blood cell transfusions in very and extremely low birthweight infants under restrictive transfusion guidelines: is exogenous erythropoietin necessary?

    PubMed Central

    Franz, A; Pohlandt, F

    2001-01-01

    OBJECTIVE—To examine the number and volume of red blood cell transfusions (RBCTs) in very and extremely low birthweight infants under restrictive red blood cell transfusion guidelines without erythropoietin administration, and to compare the results with those reported in similar infants receiving erythropoietin.
METHODS—From April 1996 to June 1999, all RBCTs given to infants with a birth weight of less than 1500 g were prospectively recorded. Data on RBCT combined with erythropoietin treatment and RBCT guidelines were extracted from four prospective randomised trials of erythropoietin for anaemia of prematurity.
RESULTS—When the restrictive RBCT guidelines were followed, the number of RBCTs and volume transfused were similar to those reported during erythropoietin administration.
CONCLUSIONS—RBCT guidelines may have a similar impact on RBCT in very low birthweight infants to the administration of erythropoietin. The effect of RBCT guidelines on RBCT frequency should be considered when evaluating the efficacy of erythropoietin administration to preterm infants.
 PMID:11207224

  5. Erythropoietin Neuroprotection in Neonatal Cardiac Surgery: A Phase I/II Safety and Efficacy Trial

    PubMed Central

    Andropoulos, Dean B.; Brady, Ken; Easley, R. Blaine; Dickerson, Heather A.; Voigt, Robert G.; Shekerdemian, Lara S.; Meador, Marcie R.; Eisenman, Carol A.; Hunter, Jill V.; Turcich, Marie; Rivera, Carlos; McKenzie, E. Dean; Heinle, Jeffrey S.; Fraser, Charles D.

    2012-01-01

    Objectives Neonates undergoing complex congenital heart surgery have a significant incidence of neurological problems. Erythropoietin has anti-apoptotic, anti-excitatory, and anti-inflammatory properties to prevent neuronal cell death in animal models, and improves neurodevelopmental outcomes in full term neonates with hypoxic ischemic encephalopathy. We designed a prospective phase I/II trial of erythropoietin neuroprotection in neonatal cardiac surgery to assess safety, and indicate efficacy. Methods Neonates undergoing surgery for D-transposition of the great vessels, hypoplastic left heart syndrome, or aortic arch reconstruction were randomized to 3 perioperative doses of erythropoietin, or placebo. Neurodevelopmental testing with Bayley Scales of Infant and Toddler Development III was performed at age 12 months. Results 59 patients received study drug. Safety profile, including MRI brain injury, clinical events, and death, was not different between groups. 3 patients in each group died. 42 patients (22 erythropoietin, 20 placebo, 79% of survivors) returned for 12-month follow-up. The mean Cognitive Scores were erythropoietin, 101.1 ± 13.6, placebo, 106.3 ± 10.8 (p=0.19); Language Scores were erythropoietin 88.5 ± 12.8, placebo 92.4 ± 12.4 (p=0.33); and Motor Scores were erythropoietin 89.9 ± 12.3, placebo 92.6 ± 14.1, (p=0.51). Conclusions Safety profile for erythropoietin administration was not different than placebo. Neurodevelopmental outcomes were not different between groups, however this pilot study was not powered to definitively address this outcome. Lessons learned from the current study suggest optimized study design features for a larger prospective trial to definitively address the utility of erythropoietin for neuroprotection in this population. PMID:23102686

  6. Erythropoietin and diabetes mellitus.

    PubMed

    Maiese, Kenneth

    2015-10-25

    Erythropoietin (EPO) is a 30.4 kDa growth factor and cytokine that governs cell proliferation, immune modulation, metabolic homeostasis, vascular function, and cytoprotection. EPO is under investigation for the treatment of variety of diseases, but appears especially suited for the treatment of disorders of metabolism that include diabetes mellitus (DM). DM and the complications of this disease impact a significant portion of the global population leading to disability and death with currently limited therapeutic options. In addition to its utility for the treatment of anemia, EPO can improve cardiac function, reduce fatigue, and improve cognition in patients with DM as well as regulate cellular energy metabolism, obesity, tissue repair and regeneration, apoptosis, and autophagy in experimental models of DM. Yet, EPO can have adverse effects that involve the vasculature system and unchecked cellular proliferation. Critical to the cytoprotective capacity and the potential for a positive clinical outcome with EPO are the control of signal transduction pathways that include protein kinase B, the mechanistic target of rapamycin, Wnt signaling, mammalian forkhead transcription factors of the O class, silent mating type information regulation 2 homolog 1 (Saccharomyces cerevisiae), and AMP activated protein kinase. Therapeutic strategies that can specifically target and control EPO and its signaling pathways hold great promise for the development of new and effective clinical treatments for DM and the complications of this disorder.

  7. Erythropoietin and diabetes mellitus

    PubMed Central

    Maiese, Kenneth

    2015-01-01

    Erythropoietin (EPO) is a 30.4 kDa growth factor and cytokine that governs cell proliferation, immune modulation, metabolic homeostasis, vascular function, and cytoprotection. EPO is under investigation for the treatment of variety of diseases, but appears especially suited for the treatment of disorders of metabolism that include diabetes mellitus (DM). DM and the complications of this disease impact a significant portion of the global population leading to disability and death with currently limited therapeutic options. In addition to its utility for the treatment of anemia, EPO can improve cardiac function, reduce fatigue, and improve cognition in patients with DM as well as regulate cellular energy metabolism, obesity, tissue repair and regeneration, apoptosis, and autophagy in experimental models of DM. Yet, EPO can have adverse effects that involve the vasculature system and unchecked cellular proliferation. Critical to the cytoprotective capacity and the potential for a positive clinical outcome with EPO are the control of signal transduction pathways that include protein kinase B, the mechanistic target of rapamycin, Wnt signaling, mammalian forkhead transcription factors of the O class, silent mating type information regulation 2 homolog 1 (Saccharomyces cerevisiae), and AMP activated protein kinase. Therapeutic strategies that can specifically target and control EPO and its signaling pathways hold great promise for the development of new and effective clinical treatments for DM and the complications of this disorder. PMID:26516410

  8. Recent Trends in Erythropoietin-mediated Neuroprotection

    PubMed Central

    McPherson, Ronald J.; Juul, Sandra E.

    2008-01-01

    Fifteen years of evidence have established that the cytokine erythropoietin offers promise as a treatment for brain injury. In particular, neonatal brain injury may be reduced or prevented by early treatment with recombinant erythropoietin. Extreme prematurity and perinatal asphyxia are common conditions associated with poor neurodevelopmental outcomes including cerebral palsy, mental retardation, hearing or visual impairment, and attention deficit hyperactivity disorder. When high doses of erythropoietin are administered systemically, a small proportion crosses the blood-brain barrier and can protect against hypoxic-ischemic brain injury. In addition to other protective effects, erythropoietin can specifically protect dopaminergic neurons. Since reduced dopamine neurotransmission contributes to attention deficit hyperactivity disorder, this condition may be amenable to erythropoietin treatment. This review focuses on the potential application of erythropoietin as a neuroprotectant with regard to neurologic complications of extreme prematurity, including attention deficit hyperactivity disorder. Recent concerns that early erythropoietin might exacerbate the pathologic neovascularization associated with retinopathy of prematurity are addressed. PMID:17936539

  9. Neuroprotective effects of recombinant human erythropoietin in the developing brain of rat after lithium-pilocarpine induced status epilepticus.

    PubMed

    Sözmen, Sule Çağlayan; Kurul, Semra Hız; Yiş, Uluç; Tuğyan, Kazım; Baykara, Başak; Yılmaz, Osman

    2012-03-01

    Status epilepticus triggers a mixture of apoptotic and necrotic cell death within the hippocampus. This neuronal loss may result in the development of epilepsy and cognitive deficits. Erythropoietin mediates a number of biological actions within the central nervous system and has been shown to be neuroprotective. In the present study, we investigated the effects of recombinant human erythropoietin on hippocampus of rat after lithium-pilocarpine induced status epilepticus. Twenty-one dam reared Wistar male rats, 21-day-old were divided into three groups: control group, lithium-pilocarpine induced status epilepticus and lithium-pilocarpine induced status epilepticus and erythropoietin treated group. Erythropoietin treated group received recombinant human erythropoietin 10 U/g intraperitoneally 40 min after pilocarpine injection for 5 days. Rats were sacrificed and brain tissues were collected at 5th day of experiment. Neuronal cell death and apoptosis were evaluated. Histopathological examination showed that erythropoietin significantly decreased neuronal cell death in CA1, CA2, CA3 and dentate gyrus regions of hippocampus. It also diminished apoptosis in the CA1 and dentate gyrus regions of hippocampus. In conclusion, erythropoietin may preserve the number of neurons and decrease apoptosis in model of status epilepticus induced by lithium-pilocarpine. This experimental study suggests that erythropoietin administration may be neuroprotective in status epilepticus.

  10. The International Reference Preparation of Erythropoietin*

    PubMed Central

    Cotes, P. Mary; Bangham, D. R.

    1966-01-01

    The WHO Expert Committee on Biological Standardization authorized the National Institute for Medical Research to establish an International Reference Preparation of Erythropoietin on the basis of the results of an international collaborative assay and to define an international unit with the agreement of the participants. The material investigated was a research standard (Erythropoietin Standard B) that had been in widespread use for some time. The preparation of this material and the estimation of its potency in terms of an earlier research standard (Erythropoietin Standard A) are described. By comparisons with other preparations of erythropoietin, it was found that Erythropoietin Standard B is a suitable standard for the bioassay of many preparations of erythropoietin in current use. On the basis of the information obtained, Erythropoietin Standard B has been established as the International Reference Preparation of Erythropoietin and the International Unit for Erythropoietin has been defined as the activity contained in 1.48 mg of the International Reference Preparation. This amount has equivalent biological activity to the unit of Erythropoietin Standard A and Standard B and thus ensures the continuity of the units already in widespread use. PMID:5297809

  11. Does erythropoietin augment noise induced hearing loss?

    PubMed

    Frederiksen, Birgitte Lidegaard; Cayé-Thomasen, Per; Lund, Søren Peter; Wagner, Niels; Asal, Korhan; Olsen, Niels Vidiendal; Thomsen, Jens

    2007-01-01

    Noise-induced hearing loss may result from excessive release of glutamate, nitrogen oxide and reactive oxygen species. The effects of these factors on the inner ear may potentially be prevented or reduced by erythropoietin (EPO), as indicated by previously demonstrated neuro-protective effects of EPO upon damage to the central nervous system and the retina. This paper reports three separate trials, conducted to investigate the hypothesis that noise-induced hearing loss is prevented or reduced by erythropoietin. The trials employed three different modes of drug application, different administration time windows and different rodent species. In trial 1, guinea pigs were exposed to 110dB SPL, 4-20kHz wide band noise (WBN) for 8h. EPO was administered to the round window membrane 24h after noise exposure, either sustained by pump for a week or by single dose middle ear instillation. In trial 2, rats were exposed to 105dB SPL, 4-20kHz WBN for 8h. EPO was administered by single dose middle ear instillation 1 or 14h after noise exposure. In trial 3, rats were exposed to 105dB SPL, 4-20kHz WBN for 8 or 3x8h. EPO was injected intraperitoneally 1h before noise exposure. Oto-acoustic emissions and auditory brainstem responses (at 16kHz) were recorded before and after noise exposure in all trials. The noise exposure induced a hearing loss in all animals. In trial 1, no recovery and no improvement of hearing occurred in any treatment group. In trial 2 and 3, a partial hearing recovery was seen. However, the hearing loss of the EPO treated animals was significantly worse than controls in trial 2. In trial 3, the hearing of the EPO treated animals exposed for 3x8h was significantly worse than controls. Thus, surprisingly, the results from 2 of the 3 present trials indicate that erythropoietin may in fact augment noise-induced hearing loss. This is contradictory to the beneficial effect of EPO reported by the vast majority of studies on stressed neural tissues. EPO administration

  12. Does erythropoietin augment noise induced hearing loss?

    PubMed

    Frederiksen, Birgitte Lidegaard; Cayé-Thomasen, Per; Lund, Søren Peter; Wagner, Niels; Asal, Korhan; Olsen, Niels Vidiendal; Thomsen, Jens

    2007-01-01

    Noise-induced hearing loss may result from excessive release of glutamate, nitrogen oxide and reactive oxygen species. The effects of these factors on the inner ear may potentially be prevented or reduced by erythropoietin (EPO), as indicated by previously demonstrated neuro-protective effects of EPO upon damage to the central nervous system and the retina. This paper reports three separate trials, conducted to investigate the hypothesis that noise-induced hearing loss is prevented or reduced by erythropoietin. The trials employed three different modes of drug application, different administration time windows and different rodent species. In trial 1, guinea pigs were exposed to 110dB SPL, 4-20kHz wide band noise (WBN) for 8h. EPO was administered to the round window membrane 24h after noise exposure, either sustained by pump for a week or by single dose middle ear instillation. In trial 2, rats were exposed to 105dB SPL, 4-20kHz WBN for 8h. EPO was administered by single dose middle ear instillation 1 or 14h after noise exposure. In trial 3, rats were exposed to 105dB SPL, 4-20kHz WBN for 8 or 3x8h. EPO was injected intraperitoneally 1h before noise exposure. Oto-acoustic emissions and auditory brainstem responses (at 16kHz) were recorded before and after noise exposure in all trials. The noise exposure induced a hearing loss in all animals. In trial 1, no recovery and no improvement of hearing occurred in any treatment group. In trial 2 and 3, a partial hearing recovery was seen. However, the hearing loss of the EPO treated animals was significantly worse than controls in trial 2. In trial 3, the hearing of the EPO treated animals exposed for 3x8h was significantly worse than controls. Thus, surprisingly, the results from 2 of the 3 present trials indicate that erythropoietin may in fact augment noise-induced hearing loss. This is contradictory to the beneficial effect of EPO reported by the vast majority of studies on stressed neural tissues. EPO administration

  13. Reevaluation of erythropoietin production by the nephron.

    PubMed

    Nagai, Takanori; Yasuoka, Yukiko; Izumi, Yuichiro; Horikawa, Kahori; Kimura, Miho; Nakayama, Yushi; Uematsu, Takayuki; Fukuyama, Takashi; Yamazaki, Taiga; Kohda, Yukimasa; Hasuike, Yukiko; Nanami, Masayoshi; Kuragano, Takahiro; Kobayashi, Noritada; Obinata, Masuo; Tomita, Kimio; Tanoue, Akito; Nakanishi, Takeshi; Kawahara, Katsumasa; Nonoguchi, Hiroshi

    2014-06-27

    Erythropoietin production has been reported to occur in the peritubular interstitial fibroblasts in the kidney. Since the erythropoietin production in the nephron is controversial, we reevaluated the erythropoietin production in the kidney. We examined mRNA expressions of erythropoietin and HIF PHD2 using high-sensitive in situ hybridization system (ISH) and protein expression of HIF PHD2 using immunohistochemistry in the kidney. We further investigated the mechanism of erythropoietin production by hypoxia in vitro using human liver hepatocell (HepG2) and rat intercalated cell line (IN-IC cells). ISH in mice showed mRNA expression of erythropoietin in proximal convoluted tubules (PCTs), distal convoluted tubules (DCTs) and cortical collecting ducts (CCDs) but not in the peritubular cells under normal conditions. Hypoxia induced mRNA expression of erythropoietin largely in peritubular cells and slightly in PCTs, DCTs, and CCDs. Double staining with AQP3 or AE1 indicated that erythropoietin mRNA expresses mainly in β-intercalated or non α/non β-intercalated cells of the collecting ducts. Immunohistochemistry in rat showed the expression of HIF PHD2 in the collecting ducts and peritubular cells and its increase by anemia in peritubular cells. In IN-IC cells, hypoxia increased mRNA expression of erythropoietin, erythropoietin concentration in the medium and protein expression of HIF PHD2. These data suggest that erythropoietin is produced by the cortical nephrons mainly in the intercalated cells, but not in the peritubular cells, in normal hematopoietic condition and by mainly peritubular cells in hypoxia, suggesting the different regulation mechanism between the nephrons and peritubular cells.

  14. Reevaluation of erythropoietin production by the nephron.

    PubMed

    Nagai, Takanori; Yasuoka, Yukiko; Izumi, Yuichiro; Horikawa, Kahori; Kimura, Miho; Nakayama, Yushi; Uematsu, Takayuki; Fukuyama, Takashi; Yamazaki, Taiga; Kohda, Yukimasa; Hasuike, Yukiko; Nanami, Masayoshi; Kuragano, Takahiro; Kobayashi, Noritada; Obinata, Masuo; Tomita, Kimio; Tanoue, Akito; Nakanishi, Takeshi; Kawahara, Katsumasa; Nonoguchi, Hiroshi

    2014-06-27

    Erythropoietin production has been reported to occur in the peritubular interstitial fibroblasts in the kidney. Since the erythropoietin production in the nephron is controversial, we reevaluated the erythropoietin production in the kidney. We examined mRNA expressions of erythropoietin and HIF PHD2 using high-sensitive in situ hybridization system (ISH) and protein expression of HIF PHD2 using immunohistochemistry in the kidney. We further investigated the mechanism of erythropoietin production by hypoxia in vitro using human liver hepatocell (HepG2) and rat intercalated cell line (IN-IC cells). ISH in mice showed mRNA expression of erythropoietin in proximal convoluted tubules (PCTs), distal convoluted tubules (DCTs) and cortical collecting ducts (CCDs) but not in the peritubular cells under normal conditions. Hypoxia induced mRNA expression of erythropoietin largely in peritubular cells and slightly in PCTs, DCTs, and CCDs. Double staining with AQP3 or AE1 indicated that erythropoietin mRNA expresses mainly in β-intercalated or non α/non β-intercalated cells of the collecting ducts. Immunohistochemistry in rat showed the expression of HIF PHD2 in the collecting ducts and peritubular cells and its increase by anemia in peritubular cells. In IN-IC cells, hypoxia increased mRNA expression of erythropoietin, erythropoietin concentration in the medium and protein expression of HIF PHD2. These data suggest that erythropoietin is produced by the cortical nephrons mainly in the intercalated cells, but not in the peritubular cells, in normal hematopoietic condition and by mainly peritubular cells in hypoxia, suggesting the different regulation mechanism between the nephrons and peritubular cells. PMID:24832733

  15. Therapeutic strategy of erythropoietin in neurological disorders.

    PubMed

    Liu, Xiang-Bao; Wang, Jiang-An; Yu, Shan Ping; Keogh, Christine L; Wei, Ling

    2008-06-01

    Erythropoietin (EPO) was first identified as a hematopoietic cytokine that stimulates proliferation and differentiation of erythroid progenitor cells and was approved by the Food and Drug Administration as a treatment for chronic renal disease patients with anemia. In neural tissues, EPO is working via EPO receptors and induces non-hematopoietic effects. Recent studies have demonstrated that EPO exerts therapeutic potentials on neurological disorders such as ischemic stroke, intracerebral hemorrhage, subarachnoid hemorrhage, traumatic brain injury, and Parkinson's disease. EPO treatment has been shown to reduce the ischemic infarct and hemorrhage volume, decrease neuronal death including apoptosis, and improve survival rates in animal models. The mechanism of EPO action in neurological disorders involves neuroprotection and promotion of neurogenesis and angiogenesis. Clinical trials of EPO treatments in neurological diseases have accumulated positive results. In stroke patients, EPO treatment may reduce infarct volume and improve functional outcomes. EPO administration has proven safe in animal studies and adult human patients, although safety and efficacy data in neonates and infants are incomplete and long-term multi-center patient evaluations are necessary. Available information suggests that EPO is a promising therapeutic drug for the treatment of neurological diseases.

  16. Recombinant erythropoietin in clinical practice

    PubMed Central

    Ng, T; Marx, G; Littlewood, T; Macdougall, I

    2003-01-01

    The introduction of recombinant human erythropoietin (RHuEPO) has revolutionised the treatment of patients with anaemia of chronic renal disease. Clinical studies have demonstrated that RHuEPO is also useful in various non-uraemic conditions including haematological and oncological disorders, prematurity, HIV infection, and perioperative therapies. Besides highlighting both the historical and functional aspects of RHuEPO, this review discusses the applications of RHuEPO in clinical practice and the potential problems of RHuEPO treatment. PMID:12897214

  17. Erythropoietin: still on the neuroprotection road

    PubMed Central

    del Barco, Diana García; Coro-Antich, Rosa M.

    2012-01-01

    Acute stroke is one of the major causes of death and disabilities. Since the 1980s many clinical studies have been conducted to evaluate neuroprotective approaches to treat this important brain vascular event. However, to date the only drug approved (recombinant tissue plasminogen activator [rtPA]) represents a thrombolytic, nonneuroprotective approach. An important neuroprotective strategy is based on erythropoietin (EPO). Exogenously administered EPO exhibits neuroprotective effects in numerous animal models, through the activation of anti-apoptotic, anti-oxidant and anti-inflammatory pathways as well as through the stimulation of angiogenic and neurogenic events. The capability of EPO to cross the blood–brain barrier after systemic administration and its effective therapeutic window are advantages for human acute stroke therapy. However, a multicenter stroke trial where recombinant human EPO (rhEPO) was combined with rtPA had negative outcomes. The present paper reviews the EPO neuroprotective strategy and its mechanisms in ischemic stroke and in other human nervous system diseases. PMID:22590480

  18. Downstream modulation of extrinsic apoptotic pathway in streptozotocin-induced Alzheimer's dementia in rats: Erythropoietin versus curcumin.

    PubMed

    Samy, Doaa M; Ismail, Cherine A; Nassra, Rasha A; Zeitoun, Teshreen M; Nomair, Azhar M

    2016-01-01

    Erythropoietin and curcumin showed promising neuroprotective effects in various models of Alzheimer's dementia. This study was designed to compare the beneficial effects of erythropoietin and/or curcumin in intracerebro-ventricular (ICV) streptozotocin-induced Alzheimer's like disease in rats. Rats received ICV injection of either saline (control, n=8 rats), or streptozotocin. Three weeks following surgery, streptozotocin-injected rats were assigned into 4 groups (8 rats each); vehicle, curcumin (80mg/kg/day, orally), erythropoietin (500 IU/kg every other day, intraperitoneally) and combined (curcumin and erythropoietin)-treated groups. After 3 months of treatment, rats were subjected to neurobehavioral testing, and then killed for biochemical and histological assessment of hippocampus. Fas ligand protein and caspase-8 activity as mediators of extrinsic apoptotic pathway, oxidative stress markers (malondialdehyde and reduced glutathione) and β-amyloid (1-40 and 1-42) peptides were measured. The results showed that administration of erythropoietin suppressed extrinsic apoptosis better than curcumin, while curcumin was more effective in combating oxidative stress in ICV-streptozotocin injected rats. Both erythropoietin and curcumin treatments (individually or combined) equally reduced the hippocampal β-amyloid accumulation and improved cognitive impairment in Morris water maze and passive avoidance tasks. The combined treatment was the most effective in ameliorating apoptosis and oxidative stress rather than behavioral responses or β-amyloid burden. In conclusion, ICV-streptozotocin-induced Alzheimer's dementia activates hippocampal Fas ligand-mediated apoptosis, which could be reduced by erythropoietin and/or curcumin treatment. Curcumin supplementation alone could ameliorate cognitive deficits and reverse biochemical alterations in ICV-streptozotocin Alzheimer's rat model without the hazardous polycythemic effect of long-term erythropoietin injection.

  19. Soluble Erythropoietin Receptor Contributes to Erythropoietin Resistance in End-Stage Renal Disease

    PubMed Central

    Tamez, Hector; Yuan, Hai-Tao; Karumanchi, S. Ananth; Thadhani, Ravi

    2010-01-01

    Background Erythropoietin is a growth factor commonly used to manage anemia in patients with chronic kidney disease. A significant clinical challenge is relative resistance to erythropoietin, which leads to use of successively higher erythropoietin doses, failure to achieve target hemoglobin levels, and increased risk of adverse outcomes. Erythropoietin acts through the erythropoietin receptor (EpoR) present in erythroblasts. Alternative mRNA splicing produces a soluble form of EpoR (sEpoR) found in human blood, however its role in anemia is not known. Methods and Findings Using archived serum samples obtained from subjects with end stage kidney disease we show that sEpoR is detectable as a 27kDa protein in the serum of dialysis patients, and that higher serum sEpoR levels correlate with increased erythropoietin requirements. Soluble EpoR inhibits erythropoietin mediated signal transducer and activator of transcription 5 (Stat5) phosphorylation in cell lines expressing EpoR. Importantly, we demonstrate that serum from patients with elevated sEpoR levels blocks this phosphorylation in ex vivo studies. Finally, we show that sEpoR is increased in the supernatant of a human erythroleukaemia cell line when stimulated by inflammatory mediators such as interleukin-6 and tumor necrosis factor alpha implying a link between inflammation and erythropoietin resistance. Conclusions These observations suggest that sEpoR levels may contribute to erythropoietin resistance in end stage renal disease, and that sEpoR production may be mediated by pro-inflammatory cytokines. PMID:20169072

  20. Symmetric signaling by an asymmetric 1 erythropoietin: 2 erythropoietin receptor complex.

    PubMed

    Zhang, Yingxin L; Radhakrishnan, Mala L; Lu, Xiaohui; Gross, Alec W; Tidor, Bruce; Lodish, Harvey F

    2009-01-30

    Via sites 1 and 2, erythropoietin binds asymmetrically to two identical receptor monomers, although it is unclear how asymmetry affects receptor activation and signaling. Here we report the design and validation of two mutant erythropoietin receptors that probe the role of individual members of the receptor dimer by selectively binding either site 1 or site 2 on erythropoietin. Ba/F3 cells expressing either mutant receptor do not respond to erythropoietin, but cells co-expressing both receptors respond to erythropoietin by proliferation and activation of the JAK2-Stat5 pathway. A truncated receptor with only one cytosolic tyrosine (Y343) is sufficient for signaling in response to erythropoietin, regardless of the monomer on which it is located. Similarly, only one receptor in the dimer needs a juxtamembrane hydrophobic L253 or W258 residue, essential for JAK2 activation. We conclude that despite asymmetry in the ligand-receptor interaction, both sides are competent for signaling, and appear to signal equally.

  1. Caveat Oncologist: Clinical Findings and Consequences of Distributing Counterfeit Erythropoietin in the United States

    PubMed Central

    Qureshi, Zaina P.; Norris, LeAnn; Sartor, Oliver; McKoy, June M.; Armstrong, John; Raisch, Dennis W.; Garg, Vishvas; Stafkey-Mailey, Dana; Bennett, Charles Lee

    2012-01-01

    Purpose: Counterfeit pharmaceuticals pose risks domestically. Because of their cost, cancer pharmaceuticals are vulnerable. We review findings from a domestic counterfeiting episode involving erythropoietin and outline anticounterfeiting recommendations for policy makers, patients, and health care professionals. Materials and Methods: Information was obtained on patients who received counterfeit erythropoietin, its distribution, and criminal investigations into counterfeiting networks. Interview sources included a physician, an attorney, employees of the Florida Department of Health and Human Services and the US Food and Drug Administration's (FDA) Office of Criminal Investigation, manufacturers, and wholesalers. Other sources included the book “Dangerous Doses,” LexisNexis (search terms “counterfeit” and “erythropoietin”) and the FDA database. Results: Counterfeit product consisted of 2,000 U vials with counterfeit labels denoting 40,000 U. The counterfeiters, in collaboration with a Miami pharmacy, purchased 110,000 erythropoietin 2,000 U vials and affixed counterfeit labels to each vial. Products were then sold via the pharmaceutical “gray market” to wholesalers, then pharmacy chains. Investigations by Florida government officials implicated 17 persons, all of whom were found guilty of trafficking in counterfeit pharmaceuticals. Despite the large size of the operation, the FDA received reports of only 12 patients who had received counterfeit erythropoietin and detailed information for only two individuals. A 17-year-old liver transplant recipient and a 61-year-old patient with breast cancer experienced loss of efficacy after receiving counterfeit erythropoietin. Conclusion: Wider use of FDA anticounterfeit initiatives, limiting pharmaceutical suppliers to reputable distributors, and educating providers and patients about signs of counterfeit drugs can improve the safety of cancer pharmaceuticals. PMID:23077434

  2. Antiapoptotic properties of recombinant human erythropoietin protects against tubular cyclosporine toxicity.

    PubMed

    Pallet, Nicolas; Bouvier, Nicolas; Legendre, Christophe; Beaune, Philippe; Thervet, Eric; Choukroun, Gabriel; Martinez, Frank

    2010-01-01

    During the early post transplant period, the tubular epithelium is the main target of injuries including ischemia reperfusion and toxicity effects from calcineurin inhibitors. Taking into account the tissue protective effects of erythropoietin mediated through its antiapoptotic properties, we tested whether administration of recombinant human erythropoietin protects against acute cyclosporine nephrotoxicity. Four groups of five rats were intraperitoneally treated over 28 days with 100UI/Kg/48h Epoetin beta (15mg/kg/day CsA diluted in olive oil, 100UI/Kg/48h Epoetin beta+15mg/kg/day CsA, or olive oil. Histological changes due to tubular necrosis were evaluated with Masson'Trichrome staining. Apoptotic nuclei in kidneys were detected using the Terminal deoxynucleotidyl Transferase Biotin-dUTP Nick End Labeling (TUNEL) method. Phospho-Akt, Akt, cleaved caspase 3 and non cleaved caspase 3 expression were evaluated using immunblotting. We demonstrate that recombinant human erythropoietin (epoetin beta) improves renal function and protects against acute tubular injury. Our data suggest that this nephroprotective effect is mediated by Akt activation and inhibition of tubular apoptosis. Indeed, western blotting analysis of caspase 3 cleavage and Akt phosphorylation demonstrates that rhEPO activate Akt signaling and inhibits caspase 3 cleavage induced by CsA. TUNEL staining confirms that rhEPO inhibits CsA-induced tubular apoptosis. In conclusion, we describe here a new potential target of recombinant human erythropoietin and our results provide an interesting framework for further nephroprotective therapies based on recombinant human erythropoietin.

  3. Development of a new radioimmunoassay for erythropoietin using recombinant erythropoietin

    SciTech Connect

    Mason-Garcia, M.; Beckman, B.S.; Brookins, J.W.; Powell, J.S.; Lanham, W.; Blaisdell, S.; Keay, L.; Li, S.C.; Fisher, J.W. )

    1990-11-01

    The development of a 24 hour radioimmunoassay for erythropoietin (EPO) using EPO derived from recombinant DNA as both immunogen and ligand is described in the present paper. Mixed breed rabbits immunized with 10 micrograms/kg of EPO derived from a stably transfected cell line (MD) produced antibodies to EPO with high titer (up to 1:896,000 final dilution in the tube), high affinity (8.4 x 10(11) liter/M), and good specificity. Purified EPO from the above source or from AmGen Biologicals (AG) were successfully radioiodinated with the chloramine-T method and used as ligand in the radioimmunoassay. Standard dose-response curves prepared with EPO from both commercial sources were not significantly different and showed a sensitivity of 0.75 to 0.96 mU/tube. The dose-response curves in both systems also showed parallelism with serially diluted serum from a patient with aplastic anemia. Within-assay and between-assay precision were determined by assaying multiple replicates of a serum pool. Recovery of exogenous EPO added to a serum pool averaged 97% for both systems. The range of normal human serum EPO was determined by assaying the sera of 153 hematologically-normal adult subjects and was found to be 1.1 to 27.3 mU/ml for MD EPO and 0.5 to 16.7 mU/ml for AG EPO. Sera from several patients with hematologic abnormalities were also assayed, including those of 36 patients with anemia of end-stage renal disease (mean +/- SEM, 29.5 +/- 4.0 mU/ml; P less than 0.01). In conclusion, this new, more rapid and sensitive radioimmunoassay system can be used to measure EPO levels in sera from normal human subjects and patients with several types of anemia, and should also be very useful in therapeutic drug monitoring of patients receiving EPO from various commercial sources.

  4. Tract-based spatial statistics to assess the neuroprotective effect of early erythropoietin on white matter development in preterm infants.

    PubMed

    O'Gorman, Ruth L; Bucher, Hans U; Held, Ulrike; Koller, Brigitte M; Hüppi, Petra S; Hagmann, Cornelia F

    2015-02-01

    infants treated with recombinant human erythropoietin than in those treated with placebo (P < 0.001). We conclude that early recombinant human erythropoietin administration improves white matter development in preterm infants assessed by diffusion tensor imaging and tract-based spatial statistics. PMID:25534356

  5. Immunomodulatory action of human recombinant erythropoietin in man.

    PubMed

    Imiela, J; Korczak-Kowalska, G; Małecki, R; Nowaczyk, M; Stepień-Sopniewska, B; Górski, A

    1993-03-01

    Recent findings suggest that recombinant human erythropoietin (rhEpo) may have an immunomodulating action. We have studied the in vitro and in vivo effects of rhEpo on immune functions in man. Low pharmacological concentrations of the hormone inhibit T-cell activation and proliferation, while higher ones are without that effect. The same Epo concentrations inhibit mitogen- and alloantigen-driven B-cell differentiation and immunoglobulin synthesis and, to a lesser extent, B-cell proliferation. In vivo treatment with rhEpo causes an initial inhibition of T- and B-cell proliferation, but with prolonged administration improved responsiveness is observed. Our data support the notion that rhEpo can regulate immune functions, a fact of potential clinical application. PMID:8514337

  6. The effect of erythropoietin on serum uric acid levels during renal ischemia reperfusion injury in rats

    PubMed Central

    Tsompos, Constantinos; Panoulis, Constantinos; Toutouzas, Konstantinos; Zografos, George; Papalois, Apostolos

    2014-01-01

    Objective: The aim of this experimental study was to assess the effect of erythropoietin on a rat model, particularly under a renal ischemia reperfusion protocol. The beneficial or lack of effects of that molecule on the excreted renal product of serum uric acid were studied biochemically. Material and methods: Forty rats were used with a mean weight of 247.7 gr. Serum uric acid levels were measured measured at 60 min after reperfusion (Groups A and C) and at 120 min after reperfusion (groups B and D). Results: 1) Erythropoietin administration non-significantly decreased the serum uric acid levels non-significantly by 0.02 mg/dL [−0.2415423 mg/dL-0.2015423 mg/dL] (p=0.8560), in accordance with the paired t-test (p=0.8438). Reperfusion time non-significantly increased the serum uric acid levels non-significantly by 0.17 mg/dL [−0.0444933 mg/dL-0.3844933 mg/dL] (p=0.1169), in accordance with the paired t-test (p=0.1648). 3) The interaction of erythropoietin administration and reperfusion time non-significantly increased the serum uric acid levels non-significantly by 0.1 mg/dL [−0.0295564 mg/dL-0.2295564 mg/dL] (p=0.1264). Conclusion: Erythropoietin administration, reperfusion time and their interaction have no significant short-term alterations on serum uric acid levels. Conclusions cannot be extracted by non-significant p-values within 2 hours. Obviously, longer study times may permit safer results. PMID:26328161

  7. DNA methyltransferase inhibition restores erythropoietin production in fibrotic murine kidneys.

    PubMed

    Chang, Yu-Ting; Yang, Ching-Chin; Pan, Szu-Yu; Chou, Yu-Hsiang; Chang, Fan-Chi; Lai, Chun-Fu; Tsai, Ming-Hsuan; Hsu, Huan-Lun; Lin, Ching-Hung; Chiang, Wen-Chih; Wu, Ming-Shiou; Chu, Tzong-Shinn; Chen, Yung-Ming; Lin, Shuei-Liong

    2016-02-01

    Renal erythropoietin-producing cells (REPCs) remain in the kidneys of patients with chronic kidney disease, but these cells do not produce sufficient erythropoietin in response to hypoxic stimuli. Treatment with HIF stabilizers rescues erythropoietin production in these cells, but the mechanisms underlying the decreased response of REPCs in fibrotic kidneys to anemic stimulation remain elusive. Here, we show that fibroblast-like FOXD1+ progenitor-derived kidney pericytes, which are characterized by the expression of α1 type I collagen and PDGFRβ, produce erythropoietin through HIF2α regulation but that production is repressed when these cells differentiate into myofibroblasts. DNA methyltransferases and erythropoietin hypermethylation are upregulated in myofibroblasts. Exposure of myofibroblasts to nanomolar concentrations of the demethylating agent 5-azacytidine increased basal expression and hypoxic induction of erythropoietin. Mechanistically, the profibrotic factor TGF-β1 induced hypermethylation and repression of erythropoietin in pericytes; these effects were prevented by 5-azacytidine treatment. These findings shed light on the molecular mechanisms underlying erythropoietin repression in kidney myofibroblasts and demonstrate that clinically relevant, nontoxic doses of 5-azacytidine can restore erythropoietin production and ameliorate anemia in the setting of kidney fibrosis in mice.

  8. DNA methyltransferase inhibition restores erythropoietin production in fibrotic murine kidneys.

    PubMed

    Chang, Yu-Ting; Yang, Ching-Chin; Pan, Szu-Yu; Chou, Yu-Hsiang; Chang, Fan-Chi; Lai, Chun-Fu; Tsai, Ming-Hsuan; Hsu, Huan-Lun; Lin, Ching-Hung; Chiang, Wen-Chih; Wu, Ming-Shiou; Chu, Tzong-Shinn; Chen, Yung-Ming; Lin, Shuei-Liong

    2016-02-01

    Renal erythropoietin-producing cells (REPCs) remain in the kidneys of patients with chronic kidney disease, but these cells do not produce sufficient erythropoietin in response to hypoxic stimuli. Treatment with HIF stabilizers rescues erythropoietin production in these cells, but the mechanisms underlying the decreased response of REPCs in fibrotic kidneys to anemic stimulation remain elusive. Here, we show that fibroblast-like FOXD1+ progenitor-derived kidney pericytes, which are characterized by the expression of α1 type I collagen and PDGFRβ, produce erythropoietin through HIF2α regulation but that production is repressed when these cells differentiate into myofibroblasts. DNA methyltransferases and erythropoietin hypermethylation are upregulated in myofibroblasts. Exposure of myofibroblasts to nanomolar concentrations of the demethylating agent 5-azacytidine increased basal expression and hypoxic induction of erythropoietin. Mechanistically, the profibrotic factor TGF-β1 induced hypermethylation and repression of erythropoietin in pericytes; these effects were prevented by 5-azacytidine treatment. These findings shed light on the molecular mechanisms underlying erythropoietin repression in kidney myofibroblasts and demonstrate that clinically relevant, nontoxic doses of 5-azacytidine can restore erythropoietin production and ameliorate anemia in the setting of kidney fibrosis in mice. PMID:26731474

  9. Impact of an Interleukin-1 Receptor Antagonist and Erythropoietin on Experimental Myocardial Ischemia/Reperfusion Injury

    PubMed Central

    Grothusen, Christina; Hagemann, Angelika; Attmann, Tim; Braesen, Jan; Broch, Ole; Cremer, Jochen; Schoettler, Jan

    2012-01-01

    Background. Revascularization of infarcted myocardium results in release of inflammatory cytokines mediating myocardial reperfusion injury and heart failure. Blockage of inflammatory pathways dampens myocardial injury and reduces infarct size. We compared the impact of the interleukin-1 receptor antagonist Anakinra and erythropoietin on myocardial ischemia/reperfusion injury. In contrast to others, we hypothesized that drug administration prior to reperfusion reduces myocardial damage. Methods and Results. 12–15 week-old Lewis rats were subjected to myocardial ischemia by a 1 hr occlusion of the left anterior descending coronary artery. After 15 min of ischemia, a single shot of Anakinra (2 mg/kg body weight (bw)) or erythropoietin (5000 IE/kg bw) was administered intravenously. In contrast to erythropoietin, Anakinra decreased infarct size (P < 0.05, N = 4/group) and troponin T levels (P < 0.05, N = 4/group). Conclusion. One-time intravenous administration of Anakinra prior to myocardial reperfusion reduces infarct size in experimental ischemia/reperfusion injury. Thus, Anakinra may represent a treatment option in myocardial infarction prior to revascularization. PMID:22649318

  10. ERYTHROPOIETIN EFFECTS ON FETAL MOUSE ERYTHROID CELLS

    PubMed Central

    Chui, David H. K.; Djaldetti, Meir; Marks, Paul A.; Rifkind, Richard A.

    1971-01-01

    The effect of the hormone, erythropoietin, on cultures of erythroblasts derived from the livers of fetal C57BL/6J mice was examined. An increase both in the content and in the rate of synthesis of normal adult mouse globin chains was detected in hormone-treated cultures. The rate of protein synthesis by individual erythroblasts does not increase in response to the hormone, whereas the absolute number of hemoglobin-synthesizing cells does increase and accounts for the observed stimulation of hemoglobin synthesis. The principal effect of erythropoietin appears to be upon the population of immature erythroid precursor cells which persists in the presence of the hormone, the cells maintaining their ability to replicate, and their capacity to differentiate into hemoglobinizing erythroblasts. In the absence of hormone, already committed erythroblasts continue their development, but erythropoiesis is not sustained. PMID:5128349

  11. Pure red cell aplasia secondary to treatment with erythropoietin.

    PubMed

    Locatelli, Francesco; Del Vecchio, Lucia

    2003-01-01

    Pure red cell aplasia (PRCA) is a rare condition defined as severe anemia secondary to the virtual absence of red blood cell precursors in the bone marrow. In the setting of patients treated with rHuEPO, the disease is generated by epoetin-induced antibodies that neutralise all the exogenous rHuEPO and cross-react with endogenous erythropoietin. As a result, serum erythropoietin levels are undetectable and erythropoiesis becomes ineffective. Only 4 cases of PRCA associated with rh-EPO have been reported before 1998. Thereafter, a sharp increase in the incidence of this rare condition has been reported, mainly associated with epoetin alpha use outside the United States. A number of possible mechanisms leading to PRCA development have been identified. Among these, modification of drug formulation and down stream processing probably has had a major role. Indeed, in 1998 the formulation of epoetin alpha in Europe was modified because of the fear of the "mad cow" syndrome. However, differences in molecule structure and glycosylation among different epoetins can not be excluded. It should also be underlined that the rise in the incidence of PRCA cases has been coincident with a major shift from intravenous to subcutaneous administration of rHuEPO. The abrupt rise in the incidence of PRCA cases observed in the last few years, deserves particular attention; however, we have to balance its severity, but extreme rarity, with the high number of chronic kidney disease patients who die each year because of cardiovascular disease that could partially be reduced by anemia treatment. PMID:14696747

  12. State-of-the-art biosimilar erythropoietins in the management of renal anemia: lessons learned from Europe and implications for US nephrologists.

    PubMed

    Covic, Adrian; Abraham, Ivo

    2015-09-01

    The European Medicines Agency (EMA), under a strictly regulated pathway, has approved several biosimilar products since 2005, including biosimilar versions of the erythropoiesis-stimulating agent (ESA) epoetin alfa since 2007. Subsequent to these approvals, the use of biosimilar epoetin alfa in the management of renal anemia has grown steadily throughout Europe. With the enactment of the US Biologics Price Competition and Innovation Act of 2009, a US Food and Drug Administration regulatory approval process for biosimilars was legalized. Thus, biosimilar erythropoietin products are expected to be available for prescription in the USA by mid-decade, presumably at a price that is competitive with the originator brand-name reference products. In this paper, we describe the status of originator and biosimilar ESAs, review the clinical development and regulatory approval of biosimilar erythropoietins in Europe, and summarize relevant efficacy and safety information of biosimilar erythropoietins in relation to their reference products to provide a background for US nephrologists as they appraise biosimilar erythropoietins as treatment options for renal anemia. Key lessons learned from Europe are that (a) EMA-approved biosimilar erythropoietins have comparable efficacy and safety profiles to their reference product erythropoietin; (b) pharmacovigilance preapproval and postapproval are critical, especially with regard to immunogenicity and vascular thromboembolic events; (c) strict preapproval and postapproval requirements must guide the regulatory pathway for biosimilars; and (d) high-quality manufacturing and production processes must be established to ensure quality biosimilar products. The availability of biosimilar erythropoietins in the USA will provide nephrologists with alternative effective, and potentially more affordable, treatment options for patients with renal anemia.

  13. Erythropoietin Pathway: A Potential Target for the Treatment of Depression

    PubMed Central

    Ma, Chongyang; Cheng, Fafeng; Wang, Xueqian; Zhai, Changming; Yue, Wenchao; Lian, Yajun; Wang, Qingguo

    2016-01-01

    During the past decade, accumulating evidence from both clinical and experimental studies has indicated that erythropoietin may have antidepressant effects. In addition to the kidney and liver, many organs have been identified as secretory tissues for erythropoietin, including the brain. Its receptor is expressed in cerebral and spinal cord neurons, the hypothalamus, hippocampus, neocortex, dorsal root ganglia, nerve axons, and Schwann cells. These findings may highlight new functions for erythropoietin, which was originally considered to play a crucial role in the progress of erythroid differentiation. Erythropoietin and its receptor signaling through JAK2 activate multiple downstream signaling pathways including STAT5, PI3K/Akt, NF-κB, and MAPK. These factors may play an important role in inflammation and neuroprogression in the nervous system. This is particularly true for the hippocampus, which is possibly related to learning, memory, neurocognitive deficits and mood alterations. Thus, the influence of erythropoietin on the downstream pathways known to be involved in the treatment of depression makes the erythropoietin-related pathway an attractive target for the development of new therapeutic approaches. Focusing on erythropoietin may help us understand the pathogenic mechanisms of depression and the molecular basis of its treatment. PMID:27164096

  14. Local Erythropoietin Injection in Tibiofibular Fracture Healing

    PubMed Central

    Bakhshi, Hooman; Kazemian, Gholamhossein; Emami, Mohammad; Nemati, Ali; Karimi Yarandi, Hossein; Safdari, Farshad

    2013-01-01

    Background Erythropoietin (EPO), in addition to its function as an erythropoiesis regulator has a regenerative activity on some nonhematopoietic tissues. Animal studies have suggested a role for erythropoietin in bone healing. Objectives The present study aimed to evaluate the effects of local EPO injection in healing of tibiofibular fractures. Materials and Methods In a prospective double blind study, 60 patients with tibiofibular fracture were divided to equal EPO or placebo groups, randomly. Patients received local injection of either EPO or a placebo to the site of fracture two weeks after surgical fixation. Patients were followed by clinical and radiographic examination to determine the union rate. The period of fracture union and incidence of nonunion were compared between the two groups. Results The demographic data and types of fractures were similar in the both groups. The mean duration of the fracture union was 2.1 weeks shorter in those treated with EPO (P = 0.01). Nonunion was observed in 6 patients of the control group and 2 receiving EPO (P = 0.02). No patient experienced any adverse effect from local EPO injections. Conclusions EPO injection into the site of tibiofibular fractures may possibly accelerate healing. PMID:24350133

  15. Erythropoietin as a new therapeutic opportunity in brain inflammation and neurodegenerative diseases.

    PubMed

    Merelli, A; Czornyj, L; Lazarowski, A

    2015-01-01

    Highly expressed Erythropoietin Receptor (EPO-R) has been detected in several nonhematopoietic hypoxic cells, including cells from different brain areas in response to many different types of cell injury. In brain, hypoxia-ischemia (HI) can induce a wide spectrum of biologic responses, where inflammation and apoptosis are the main protagonists. Inflammation, as a primary brain insult, can induce a chronic hypoxic condition, producing the continuous cycle of inflammation-hypoxia that increases the apoptotic-cell number. It has also been demonstrated that administration of erythropoietin (EPO) prevented the neuronal death induced by HI, as well as the induction of lipid peroxidation in the hippocampus in a rodent model of Alzheimer's disease. Anti-apoptotic, anti-inflammatory, anti-oxidant, and/or cell-proliferative effects of EPO, have been observed in all type of cells expressing EPO-R, resulting in a potential tool for neuroprotection, neuroreparation, or neurogenesis of brain damaged areas. The nasal route is an alternative way of drugs administration that has been successfully exploited for bypassing the blood brain barrier, and subsequently delivering EPO and other molecules to central nervous system. Intranasal administration of EPO could be a new therapeutic opportunity in several brain damages that includes hypoxia, inflammation, neurodegenerative process, and apoptosis. PMID:25405533

  16. Erythropoietin stimulation decreases hepcidin expression through hematopoietic activity on bone marrow cells in mice.

    PubMed

    Sasaki, Yusuke; Noguchi-Sasaki, Mariko; Yasuno, Hideyuki; Yorozu, Keigo; Shimonaka, Yasushi

    2012-12-01

    Erythropoiesis-stimulating agents (ESA) are now central to the treatment of renal anemia and are associated with improved clinical outcomes. It is well known that erythropoietin (EPO) is a key regulator of erythropoiesis through its promotion of red blood cell production. In order to investigate the role of ESA on iron metabolism, we analyzed the regulation of the iron regulatory hormone hepcidin by ESA treatment in a bone marrow transplant model in mouse. After treating C57BL/6 mice with continuous erythropoietin receptor activator (C.E.R.A.), recombinant human epoetin-β (rhEPO), or recombinant human carbamylated epoetin-β (rhCEPO), we investigated serum hepcidin concentrations and parameters of erythropoiesis. Serum hepcidin concentrations after rhEPO treatment were analyzed in mice subjected to total body irradiation followed by bone marrow transplantation. C.E.R.A. administration caused long-term downregulation of serum hepcidin levels. Serum hepcidin levels in rhEPO-treated mice decreased significantly, whereas there was no change in rhCEPO-treated mice. The reduction in circulating hepcidin levels after rhEPO administration was not observed in irradiated mice. Finally, bone marrow transplantation recovered the response to rhEPO administration that downregulates hepcidin concentration in irradiated mice. These results indicate that ESA treatment downregulates serum hepcidin concentrations, mainly by indirect mechanisms affecting hematopoietic activity in bone marrow cells. PMID:23160767

  17. Erythropoietin Levels in Elderly Patients with Anemia of Unknown Etiology

    PubMed Central

    Sriram, Swetha; Martin, Alison; Xenocostas, Anargyros; Lazo-Langner, Alejandro

    2016-01-01

    Background In many elderly patients with anemia, a specific cause cannot be identified. This study investigates whether erythropoietin levels are inappropriately low in these cases of “anemia of unknown etiology” and whether this trend persists after accounting for confounders. Methods This study includes all anemic patients over 60 years old who had erythropoietin measured between 2005 and 2013 at a single center. Three independent reviewers used defined criteria to assign each patient’s anemia to one of ten etiologies: chronic kidney disease, iron deficiency, chronic disease, confirmed myelodysplastic syndrome (MDS), suspected MDS, vitamin B12 deficiency, folate deficiency, anemia of unknown etiology, other etiology, or multifactorial etiology. Iron deficiency anemia served as the comparison group in all analyses. We used linear regression to model the relationship between erythropoietin and the presence of each etiology, sequentially adding terms to the model to account for the hemoglobin concentration, estimated glomerular filtration rate (eGFR) and Charlson Comorbidity Index. Results A total of 570 patients met the inclusion criteria. Linear regression analysis showed that erythropoietin levels in chronic kidney disease, anemia of chronic disease and anemia of unknown etiology were lower by 48%, 46% and 27%, respectively, compared to iron deficiency anemia even after adjusting for hemoglobin, eGFR and comorbidities. Conclusions We have shown that erythropoietin levels are inappropriately low in anemia of unknown etiology, even after adjusting for confounders. This suggests that decreased erythropoietin production may play a key role in the pathogenesis of anemia of unknown etiology. PMID:27310832

  18. Human Erythropoietin Dimers with Markedly Enhanced in vivo Activity

    NASA Astrophysics Data System (ADS)

    Sytkowski, Arthur J.; Dotimas Lunn, Elizabeth; Davis, Kerry Lynn; Feldman, Laurie; Siekman, Suvia

    1998-02-01

    Human erythropoietin, a widely used and important therapeutic glycoprotein, has a relatively short plasma half-life due to clearance by glomerular filtration as well as by other mechanisms. We hypothesized that an erythropoietin species with a larger molecular size would exhibit an increased plasma half-life and, potentially, an enhanced biological activity. We now report the production of biologically active erythropoietin dimers and trimers by chemical crosslinking of the conventional monomeric form. We imparted free sulfhydryl residues to a pool of erythropoietin monomer by chemical modification. A second pool was reacted with another modifying reagent to yield monomer with male-imido groups. Upon mixing these two pools, covalently linked dimers and trimers were formed that were biologically active in vitro. The plasma half-life of erythropoietin dimers in rabbits was >24 h compared with 4 h for the monomers. Importantly, erythropoietin dimers were biologically active in vivo as shown by their ability to increase the hematocrits of mice when injected subcutaneously. In addition, the dimers exhibited >26-fold higher activity in vivo than did the monomers and were very effective after only one dose. Dimeric and other oligomeric forms of Epo may have an important role in therapy.

  19. Current perspectives on protective roles of erythropoietin in cardiovascular system: erythropoietin receptor as a novel therapeutic target.

    PubMed

    Kagaya, Yutaka; Asaumi, Yasuhide; Wang, Wanting; Takeda, Morihiko; Nakano, Makoto; Satoh, Kimio; Fukumoto, Yoshihiro; Shimokawa, Hiroaki

    2012-01-01

    Erythropoietin (EPO) is a principal regulator that promotes proliferation and terminal differentiation of erythroid progenitor cells. EPO receptors are expressed not only in hematopoietic lineage cells but also in the cardiovascular system. We performed animal experiments using transgene-rescued EPO receptor null mutant mice (EpoR-/- rescued) that express the EPO receptor exclusively in the hematopoietic cells. The results of these experiments suggest that endogenous EPO/EPO receptor system in the heart exerts cardioprotective effects against myocardial injury induced by ischemia followed by reperfusion and pressure-overload induced left ventricular dysfunction. Many animal experiments have shown that the administration of recombinant human EPO also elicits cardioprotective effects against myocardial injury induced by ischemia and reperfusion. In contrast to the promising results of these animal experiments, recent clinical trials failed to demonstrate the reduction in infarct size or improvement of cardiac function by the administration of recombinant human EPO in patients with acute myocardial infarction who underwent primary percutaneous coronary intervention. It should be tested in future clinical studies whether a relatively low dose of recombinant human EPO or its derivatives that have no erythropoietic action reduces infarct size and ameliorates cardiac dysfunction in patients with acute myocardial infarction. In this article, we review implications of anemia associated with chronic heart failure, roles of the endogenous EPO/EPO receptor system, and the effects of the administration of erythropoiesis-stimulating agents in pathologic conditions of the heart by focusing on the EPO receptor as a potential candidate of novel therapeutic targets in cardiovascular diseases.

  20. Regeneration in the nervous system with erythropoietin

    PubMed Central

    Maiese, Kenneth

    2015-01-01

    Globally, greater than 30 million individuals are afflicted with disorders of the nervous system accompanied by tens of thousands of new cases annually with limited, if any, treatment options. Erythropoietin (EPO) offers an exciting and novel therapeutic strategy to address both acute and chronic neurodegenerative disorders. EPO governs a number of critical protective and regenerative mechanisms that can impact apoptotic and autophagic programmed cell death pathways through protein kinase B (Akt), sirtuins, mammalian forkhead transcription factors, and wingless signaling. Translation of the cytoprotective pathways of EPO into clinically effective treatments for some neurodegenerative disorders has been promising, but additional work is necessary. In particular, development of new treatments with erythropoiesis-stimulating agents such as EPO brings several important challenges that involve detrimental vascular outcomes and tumorigenesis. Future work that can effectively and safely harness the complexity of the signaling pathways of EPO will be vital for the fruitful treatment of disorders of the nervous system. PMID:26549969

  1. Successful Treatment of Severe Anemia using Erythropoietin in a Jehovah Witness with Non-Hodgkin Lymphoma

    PubMed Central

    Agapidou, Alexandra; Vakalopoulou, Sofia; Papadopoulou, Theodosia; Chadjiaggelidou, Christina; Garypidou, Vasileia

    2014-01-01

    Blood transfusion many times works in a life-saving way when a patient is facing a critical situation. However, some patients, such as Jehovah’s Witnesses, may refuse their administration because it opposes to their religion beliefs. Thus, clinicians are forced to respect patients’ preferences and seek other treatments in order to overcome the obstacle of the transfusion. In 1989, recombinant human erythropoietin (rHuEPO) was approved by the United States Food and Drug Administration (FDA) for the treatment of anemia associated with chronic renal failure. This is an amino acid glycol-protein that stimulates red blood cell production in the same manner as endogenous erythropoietin. Other treatment indications approved by the FDA include anemia due to chronic kidney disease, anemia secondary to zidovudine therapy in patients with human immunodeficiency virus infection, and anemia secondary to cancer chemotherapy. The drug also has been used for many off-label indications. Many Jehovah’s Witnesses have accepted rHuEPO as a treatment option to maintain and enhance erythropoiesis. This paper reports the case of a 57-year-old Jehovah’s Witness man, who was diagnosed with severe anemia due to aggressive non Hodgkin lymphoma and refused transfusion of blood; thanks to the treatment with rHuEPO he has managed to complete chemotherapy and has survived a life threatening situation. PMID:25568760

  2. Successful Treatment of Severe Anemia using Erythropoietin in a Jehovah Witness with Non-Hodgkin Lymphoma.

    PubMed

    Agapidou, Alexandra; Vakalopoulou, Sofia; Papadopoulou, Theodosia; Chadjiaggelidou, Christina; Garypidou, Vasileia

    2014-11-19

    Blood transfusion many times works in a life-saving way when a patient is facing a critical situation. However, some patients, such as Jehovah's Witnesses, may refuse their administration because it opposes to their religion beliefs. Thus, clinicians are forced to respect patients' preferences and seek other treatments in order to overcome the obstacle of the transfusion. In 1989, recombinant human erythropoietin (rHuEPO) was approved by the United States Food and Drug Administration (FDA) for the treatment of anemia associated with chronic renal failure. This is an amino acid glycol-protein that stimulates red blood cell production in the same manner as endogenous erythropoietin. Other treatment indications approved by the FDA include anemia due to chronic kidney disease, anemia secondary to zidovudine therapy in patients with human immunodeficiency virus infection, and anemia secondary to cancer chemotherapy. The drug also has been used for many off-label indications. Many Jehovah's Witnesses have accepted rHuEPO as a treatment option to maintain and enhance erythropoiesis. This paper reports the case of a 57-year-old Jehovah's Witness man, who was diagnosed with severe anemia due to aggressive non Hodgkin lymphoma and refused transfusion of blood; thanks to the treatment with rHuEPO he has managed to complete chemotherapy and has survived a life threatening situation. PMID:25568760

  3. Pharmacological Effects of Erythropoietin and its Derivative Carbamyl erythropoietin in Cerebral White Matter Injury

    NASA Astrophysics Data System (ADS)

    Liu, Wei

    Periventricular leukomalacia (PVL) is the predominant form of brain injury in the premature infant and the most common cause of cerebral palsy, yet no therapy currently exists for this serious human disorder. As PVL often occurs in preterm infants suffering from cerebral hypoxia/ischemia with or without prior exposure to maternal-fetal infection/inflammation, we used hypoxia/ischemia with or without lipopolysaccharide (LPS) injection, to produce clinically relevant PVL-like lesions in the white matter in postnatal day six (P6) mice. We studied the white matter pathology under different conditions, such as different durations of hypoxia and different doses of LPS, to evaluate the effects of those etiological factors on neonatal white matter injury. Distinct related pathological events were investigated at different time points during the progression of PVL. We used immunohistochemistry, histological analysis, and electron microscopy (EM) to study demylination that occurs in the white matter area, which is consistent with the pathology of human PVL. Previous studies have shown that erythropoietin (EPO) and its derivative carbamylated EPO (CEPO) are neuroprotective in various experimental models of brain injury. However, none of these studies investigated their efficacy against white matter injury using appropriate animal models of PVL. We produced unilateral or bilateral white matter injury in P6 mice using unilateral carotid ligation (UCL) followed by hypoxia (6% oxygen, 35 min) or by UCL/hypoxia plus LPS injection, respectively. We administered a single intraperitoneal (i.p.) dose of EPO or CEPO (5000 IU/kg) immediately after the insult, and found both drugs to provide significant protection against white matter injury in PVL mice compared to vehicle-treated groups. In addition, EPO and CEPO treatments attenuated neurobehavioral dysfunctions in an acute manner after PVL injury. EPO and CEPO have relatively few adverse effects, and thus may be a therapeutic agent

  4. Anti-erythropoietin antibodies followed by endogenous erythropoietin production in a dialysis patient.

    PubMed

    Mattison, Parnell; Upadhyay, Kiran; Wilcox, Jennifer E; Moudgil, Asha; Silverstein, Douglas M

    2010-05-01

    We present a case of a young girl with end-stage renal disease secondary to anti-glomerular basement membrane disease who was receiving maintenance peritoneal dialysis and developed pure red cell aplasia secondary to anti-erythropoietin (EPO) antibodies. This occurred 13 months after the initiation of EPO alfa therapy for anemia. Initially, the patient required intermittent red blood cell transfusions. After immunosuppressive therapy had been initiated with corticosteroids and cyclosporine, the EPO antibody levels decreased precipitously, associated with an increased level of endogenous EPO production. For the following 6 months, the patient maintained adequate (>10 g/dL) hemoglobin levels and did not require red cell transfusions.

  5. The effects of erythropoietin signaling on telomerase regulation in non-erythroid malignant and non-malignant cells

    SciTech Connect

    Uziel, Orit; Kanfer, Gil; Beery, Einat; Yelin, Dana; Shepshelovich, Daniel; Bakhanashvili, Mary; Nordenberg, Jardena; Lahav, Meir

    2014-07-18

    Highlights: • We assumed that some of erythropoietin adverse effects may be mediated by telomerase activity. • EPO administration increased telomerase activity, cells proliferation and migration. • The inhibition of telomerase modestly repressed the proliferative effect of erythropoietin. • Telomere shortening caused by long term inhibition of the enzyme totally abolished that effect. • This effect was mediated via the Lyn–AKT axis and not by the canonical JAK2–STAT pathway. - Abstract: Treatment with erythropoietin (EPO) in several cancers is associated with decreased survival due to cancer progression. Due to the major importance of telomerase in cancer biology we hypothesized that some of these effects may be mediated through EPO effect on telomerase. For this aim we explored the possible effects of EPO on telomerase regulation, cell migration and chemosensitivity in non-erythroid malignant and non-malignant cells. Cell proliferation, telomerase activity (TA) and cell migration increased in response to EPO. EPO had no effect on cancer cells sensitivity to cisplatinum and on the cell cycle status. The inhibition of telomerase modestly repressed the proliferative effect of EPO. Telomere shortening caused by long term inhibition of the enzyme abolished the effect of EPO, suggesting that EPO effects on cancer cells are related to telomere dynamics. TA was correlated with the levels of Epo-R. The increase in TA was mediated post-translationally through the Lyn-Src and not the canonical JAK2 pathway.

  6. Erythropoietin in the treatment of carbon monoxide neurotoxicity in rat.

    PubMed

    Moallem, Seyed Adel; Mohamadpour, Amir Hooshang; Abnous, Khalil; Sankian, Mojtaba; Sadeghnia, Hamid Reza; Tsatsakis, Aristidis; Shahsavand, Shabnam

    2015-12-01

    Erythropoietin (EPO) plays a critical role in the development of the nervous system. In this study, the effects of EPO in carbon monoxide (CO) neurotoxicity were examined. Rats were exposed to 3000 ppm CO for 1 h and then different doses of EPO were administrated intraperitoneally. After 24 h, glial fibrillary acidic protein (GFAP) levels in the serum were determined and water content of brain and the extravasation of a tracer (Evans blue) were measured. Brain lipid peroxidation, myeloperoxidase activity Myelin basic protein (MBP) and BAX/BcL2 protein relative expressions were determined. Cation exchange chromatography was used to evaluate MBP alterations. Seven days after exposure, pathological assessment was performed after Klüver-Barrera staining. EPO reduced malondialdehyde levels at all doses (2500, 5000 and 10,000 u/kg). Lower doses of EPO (625, 1250, 2500 u/kg) significantly decreased the elevated serum levels of GFAP. EPO could not reduce the water content of the edematous poisoned brains. However, at 5000 and 10,000 u/kg it protected the blood brain barrier against integrity loss as a result of CO. EPO could significantly decrease the MPO activity. CO-mediated oxidative stress caused chemical alterations in MBP and EPO could partially prevent these biochemical changes. Fewer vacuoles and demyelinated fibers were found in the EPO-treated animals. EPO (5000 u/kg) could restore the MBP density. CO increased brain BAX/Bcl-2 ratio 38.78%. EPO reduced it 38.86%. These results reveal that EPO could relatively prevent different pathways of neurotoxicity by CO poisoning and thus has the potential to be used as a novel approach to manage this poisoning. PMID:26416356

  7. Erythropoietin in the treatment of carbon monoxide neurotoxicity in rat.

    PubMed

    Moallem, Seyed Adel; Mohamadpour, Amir Hooshang; Abnous, Khalil; Sankian, Mojtaba; Sadeghnia, Hamid Reza; Tsatsakis, Aristidis; Shahsavand, Shabnam

    2015-12-01

    Erythropoietin (EPO) plays a critical role in the development of the nervous system. In this study, the effects of EPO in carbon monoxide (CO) neurotoxicity were examined. Rats were exposed to 3000 ppm CO for 1 h and then different doses of EPO were administrated intraperitoneally. After 24 h, glial fibrillary acidic protein (GFAP) levels in the serum were determined and water content of brain and the extravasation of a tracer (Evans blue) were measured. Brain lipid peroxidation, myeloperoxidase activity Myelin basic protein (MBP) and BAX/BcL2 protein relative expressions were determined. Cation exchange chromatography was used to evaluate MBP alterations. Seven days after exposure, pathological assessment was performed after Klüver-Barrera staining. EPO reduced malondialdehyde levels at all doses (2500, 5000 and 10,000 u/kg). Lower doses of EPO (625, 1250, 2500 u/kg) significantly decreased the elevated serum levels of GFAP. EPO could not reduce the water content of the edematous poisoned brains. However, at 5000 and 10,000 u/kg it protected the blood brain barrier against integrity loss as a result of CO. EPO could significantly decrease the MPO activity. CO-mediated oxidative stress caused chemical alterations in MBP and EPO could partially prevent these biochemical changes. Fewer vacuoles and demyelinated fibers were found in the EPO-treated animals. EPO (5000 u/kg) could restore the MBP density. CO increased brain BAX/Bcl-2 ratio 38.78%. EPO reduced it 38.86%. These results reveal that EPO could relatively prevent different pathways of neurotoxicity by CO poisoning and thus has the potential to be used as a novel approach to manage this poisoning.

  8. Erythropoietin Restores Long-Term Neurocognitive Function Involving Mechanisms of Neuronal Plasticity in a Model of Hyperoxia-Induced Preterm Brain Injury.

    PubMed

    Hoeber, Daniela; Sifringer, Marco; van de Looij, Yohan; Herz, Josephine; Sizonenko, Stéphane V; Kempe, Karina; Serdar, Meray; Palasz, Joanna; Hadamitzky, Martin; Endesfelder, Stefanie; Fandrey, Joachim; Felderhoff-Müser, Ursula; Bendix, Ivo

    2016-01-01

    Cerebral white and grey matter injury is the leading cause of an adverse neurodevelopmental outcome in prematurely born infants. High oxygen concentrations have been shown to contribute to the pathogenesis of neonatal brain damage. Here, we focused on motor-cognitive outcome up to the adolescent and adult age in an experimental model of preterm brain injury. In search of the putative mechanisms of action we evaluated oligodendrocyte degeneration, myelination, and modulation of synaptic plasticity-related molecules. A single dose of erythropoietin (20,000 IU/kg) at the onset of hyperoxia (24 hours, 80% oxygen) in 6-day-old Wistar rats improved long-lasting neurocognitive development up to the adolescent and adult stage. Analysis of white matter structures revealed a reduction of acute oligodendrocyte degeneration. However, erythropoietin did not influence hypomyelination occurring a few days after injury or long-term microstructural white matter abnormalities detected in adult animals. Erythropoietin administration reverted hyperoxia-induced reduction of neuronal plasticity-related mRNA expression up to four months after injury. Thus, our findings highlight the importance of erythropoietin as a neuroregenerative treatment option in neonatal brain injury, leading to improved memory function in adolescent and adult rats which may be linked to increased neuronal network connectivity. PMID:27493706

  9. Erythropoietin Restores Long-Term Neurocognitive Function Involving Mechanisms of Neuronal Plasticity in a Model of Hyperoxia-Induced Preterm Brain Injury

    PubMed Central

    Sifringer, Marco; van de Looij, Yohan; Herz, Josephine; Sizonenko, Stéphane V.; Kempe, Karina; Palasz, Joanna; Hadamitzky, Martin; Fandrey, Joachim

    2016-01-01

    Cerebral white and grey matter injury is the leading cause of an adverse neurodevelopmental outcome in prematurely born infants. High oxygen concentrations have been shown to contribute to the pathogenesis of neonatal brain damage. Here, we focused on motor-cognitive outcome up to the adolescent and adult age in an experimental model of preterm brain injury. In search of the putative mechanisms of action we evaluated oligodendrocyte degeneration, myelination, and modulation of synaptic plasticity-related molecules. A single dose of erythropoietin (20,000 IU/kg) at the onset of hyperoxia (24 hours, 80% oxygen) in 6-day-old Wistar rats improved long-lasting neurocognitive development up to the adolescent and adult stage. Analysis of white matter structures revealed a reduction of acute oligodendrocyte degeneration. However, erythropoietin did not influence hypomyelination occurring a few days after injury or long-term microstructural white matter abnormalities detected in adult animals. Erythropoietin administration reverted hyperoxia-induced reduction of neuronal plasticity-related mRNA expression up to four months after injury. Thus, our findings highlight the importance of erythropoietin as a neuroregenerative treatment option in neonatal brain injury, leading to improved memory function in adolescent and adult rats which may be linked to increased neuronal network connectivity. PMID:27493706

  10. Erythropoietin and the heart: facts and perspectives.

    PubMed

    Mastromarino, Vittoria; Volpe, Massimo; Musumeci, Maria B; Autore, Camillo; Conti, Elena

    2011-01-01

    EPO (erythropoietin) has long been identified as a primary regulator of erythropoiesis. Subsequently, EPO has been recognized as playing a role in a broad variety of processes in cardiovascular pathophysiology. In particular, the tight interactions of EPO with the nitric oxide pathway, apoptosis, ischaemia, cell proliferation and platelet activation appear of great interest. Although enhanced EPO synthesis is viewed as an appropriate compensatory mechanism in the cardio-renal syndrome, which features CHF (congestive heart failure) and CRF (chronic renal failure), maladaptative excessive EPO synthesis in the advanced stages of these diseases appears to be predictive of higher mortality. Clinical trials based on the use of EPO in both heart and renal failure have so far produced contradictory results, whereas treatment targeted to restore low Hb levels appears rational and is supported by regulatory authorities. New areas for therapeutic use of EPO, such as acute coronary syndromes, are under investigation, and they are discussed in the present review together with other clinical applications in cardiovascular diseases. The revisited concept of a potential use of endogenous EPO levels as a predictor of CHF severity, as well as in the monitoring of responses to treatment, deserves appropriate investigation, as this may identify EPO as a useful biomarker in the clinical management of cardiovascular diseases. PMID:20929439

  11. Recombinant human erythropoietin and high flux haemodiafiltration.

    PubMed

    Lippi, A; Rindi, P; Baronti, R; Caprioli, R; Favilla, G; Palmarini, D; Cioni, L

    1995-01-01

    Since 1982, 32 uraemic patients were treated in our institution by high flux haemodiafiltration (H-HDF) in order to shorten significantly the dialytic treatment session. H-HDF used a high surface area filter (1.4-1.9 m2) with high hydraulic permeability (polyacrylonitrile and polysulfone), at high blood flow (450 ml/min) and high rates of reinfusion of substitution fluid (22 l/session). In this way the dialytic session was shortened to 140 +/- 19 min, maintaining a good cardiovascular stability and high dialytic efficiency (Kt/V > 1.1). Human recombinant erythropoietin rHuEpo introduced in the therapy of this group in 1987 has resulted in an improvement of renal anaemia, but also a prolongation of the time of dialytic treatment due to a decrease in the efficiency of filters. During the period of the study, the treatment time increased from 140 +/- 19 min to 168 +/- 25 min with a concomitant increase of haematocrit and haemoglobin (from 24% to 36% and from 7.9 to 10.5 g/dl, respectively). H-HDF maintains a noticeable increase in dialytic efficacy with good cardiovascular stability, but the goal of a significant reduction in the time of treatment can no longer be obtained. PMID:8524496

  12. Erythropoietin Couples Hematopoiesis with Bone Formation

    PubMed Central

    Ziegler, Anne M.; Pedersen, Elisabeth A.; Wang, Jianhua; Wang, Zhuo; Song, Junhui; Wang, Jingcheng; Lee, Clara H.; Sud, Sudha; Pienta, Kenneth J.; Krebsbach, Paul H.; Taichman, Russell S.

    2010-01-01

    Background It is well established that bleeding activates the hematopoietic system to regenerate the loss of mature blood elements. We have shown that hematopoietic stem cells (HSCs) isolated from animals challenged with an acute bleed regulate osteoblast differentiation from marrow stromal cells. This suggests that HSCs participate in bone formation where the molecular basis for this activity is the production of BMP2 and BMP6 by HSCs. Yet, what stimulates HSCs to produce BMPs is unclear. Methodology/Principal Findings In this study, we demonstrate that erythropoietin (Epo) activates Jak-Stat signaling pathways in HSCs which leads to the production of BMPs. Critically, Epo also directly activates mesenchymal cells to form osteoblasts in vitro, which in vivo leads to bone formation. Importantly, Epo first activates osteoclastogenesis which is later followed by osteoblastogenesis that is induced by either Epo directly or the expression of BMPs by HSCs to form bone. Conclusions/Significance These data for the first time demonstrate that Epo regulates the formation of bone by both direct and indirect pathways, and further demonstrates the exquisite coupling between hematopoesis and osteopoiesis in the marrow. PMID:20523730

  13. Cytoprotective effect of glutaraldehyde erythropoietin on HEK293 kidney cells after silver nanoparticle exposure

    PubMed Central

    Sooklert, Kanidta; Chattong, Supreecha; Manotham, Krissanapong; Boonwong, Chawikan; Klaharn, I-yanut; Jindatip, Depicha; Sereemaspun, Amornpun

    2016-01-01

    The toxic effects from exposure to silver nanoparticles (AgNPs), which are broadly present in many consumer products, have long raised concerns. Many studies have focused on the mechanisms of nanosilver, which cause toxicity in human cells, but little is known about prevention of this type of injury. This study investigated the in vitro effects of glutaraldehyde erythropoietin (GEPO), a cytoprotective compound derived from erythropoietin, in terms of cell protection against AgNP-induced injury. HEK293 cells were pretreated with or without GEPO before administration of AgNPs. The protective effects of GEPO in this cell line were assessed by the percentage of viable cells, alterations of cell morphology, and the proliferative capability of the cells. In addition, we assessed the role of GEPO in lowering cellular oxidative stress and regulating expression of the anti-apoptotic protein Bcl2. The results showed rescue effects on the percentage of viable and proliferative cells among GEPO pretreated cells. Pretreatment with GEPO maintained the normal cell shape and ultrastructural morphology. Moreover, GEPO reduced the generation of reactive oxygen species in cells and activated expression of Bcl2, which are the major mechanisms in protection against cellular toxicity induced by AgNPs. In conclusion, our study showed that the cytotoxic effects from exposure to AgNPs can be prevented by GEPO. PMID:26929619

  14. Neurobehavioral and cytotoxic effects of vanadium during oligodendrocyte maturation: a protective role for erythropoietin.

    PubMed

    Mustapha, Oluwaseun; Oke, Bankole; Offen, Nils; Sirén, Anna-Leena; Olopade, James

    2014-07-01

    Vanadium exposure has been known to lead to lipid peroxidation, demyelination and oligodendrocytes depletion. We investigated behaviour and glial reactions in juvenile mice after early neonatal exposure to vanadium, and examined the direct effects of vanadium in oligodendrocyte progenitor cultures from embryonic mice. Neonatal pups exposed to vanadium via lactation for 15 and 22 days all had lower body weights. Behavioural tests showed in most instances a reduction in locomotor activity and negative geotaxis. Brain analyses revealed astrocytic activation and demyelination in the vanadium exposed groups compared to the controls. In cell culture, exposure of oligodendrocytes to 300 μM sodium metavanadate significantly increased cell death. Expression of the oligodendrocyte specific proteins, 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) and oligodendrocyte specific protein (OSP/Claudin) were reduced upon vanadium treatment while simultaneous administration of erythropoietin (EPO; 4-12 U/ml) counteracted vanadium-toxicity. The data suggest that oligodendrocyte damage may explain the increased vulnerability of the juvenile brain to vanadium and support a potential for erythropoietin as a protective agent against vanadium-toxicity during perinatal brain development and maturation. PMID:24927405

  15. Neurobehavioral and cytotoxic effects of vanadium during oligodendrocyte maturation: a protective role for erythropoietin.

    PubMed

    Mustapha, Oluwaseun; Oke, Bankole; Offen, Nils; Sirén, Anna-Leena; Olopade, James

    2014-07-01

    Vanadium exposure has been known to lead to lipid peroxidation, demyelination and oligodendrocytes depletion. We investigated behaviour and glial reactions in juvenile mice after early neonatal exposure to vanadium, and examined the direct effects of vanadium in oligodendrocyte progenitor cultures from embryonic mice. Neonatal pups exposed to vanadium via lactation for 15 and 22 days all had lower body weights. Behavioural tests showed in most instances a reduction in locomotor activity and negative geotaxis. Brain analyses revealed astrocytic activation and demyelination in the vanadium exposed groups compared to the controls. In cell culture, exposure of oligodendrocytes to 300 μM sodium metavanadate significantly increased cell death. Expression of the oligodendrocyte specific proteins, 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) and oligodendrocyte specific protein (OSP/Claudin) were reduced upon vanadium treatment while simultaneous administration of erythropoietin (EPO; 4-12 U/ml) counteracted vanadium-toxicity. The data suggest that oligodendrocyte damage may explain the increased vulnerability of the juvenile brain to vanadium and support a potential for erythropoietin as a protective agent against vanadium-toxicity during perinatal brain development and maturation.

  16. Hydrogel delivery of erythropoietin to the brain for endogenous stem cell stimulation after stroke injury.

    PubMed

    Wang, Yuanfei; Cooke, Michael J; Morshead, Cindi M; Shoichet, Molly S

    2012-03-01

    Drug delivery to the brain is challenging because systemic delivery requires high doses to achieve diffusion across the blood-brain barrier and often results in systemic toxicity. Intracerebroventricular implantation of a minipump/catheter system provides local delivery, yet results in brain tissue damage and can be prone to infection. An alternate local delivery strategy, epi-cortical delivery, releases the biomolecule directly to the brain while causing minimal tissue disruption. We pursued this strategy with a hyaluronan/methyl cellulose (HAMC) hydrogel for the local release of erythropoietin to induce endogenous neural stem and progenitor cells of the subventricular zone to promote repair after stroke injury in the mouse brain. Erythropoeitin promotes neurogenesis when delivered intraventricularly, thereby making it an ideal biomolecule with which to test this new epi-cortical delivery strategy. We investigated HAMC in terms of the host tissue response and the diffusion of erythropoeitin therefrom in the stroke-injured brain for neural repair. Erythropoietin delivered from HAMC at 4 and 11 days post-stroke resulted in attenuated inflammatory response, reduced stroke cavity size, increased number of both neurons in the peri-infarct region and migratory neuroblasts in the subventricular zone, and decreased apoptosis in both the subventricular zone and the injured cortex. We demonstrate that HAMC-mediated epi-cortical administration is promising for minimally invasive delivery of erythropoeitin to the brain. PMID:22217804

  17. Cytoprotective effect of glutaraldehyde erythropoietin on HEK293 kidney cells after silver nanoparticle exposure.

    PubMed

    Sooklert, Kanidta; Chattong, Supreecha; Manotham, Krissanapong; Boonwong, Chawikan; Klaharn, I-yanut; Jindatip, Depicha; Sereemaspun, Amornpun

    2016-01-01

    The toxic effects from exposure to silver nanoparticles (AgNPs), which are broadly present in many consumer products, have long raised concerns. Many studies have focused on the mechanisms of nanosilver, which cause toxicity in human cells, but little is known about prevention of this type of injury. This study investigated the in vitro effects of glutaraldehyde erythropoietin (GEPO), a cytoprotective compound derived from erythropoietin, in terms of cell protection against AgNP-induced injury. HEK293 cells were pretreated with or without GEPO before administration of AgNPs. The protective effects of GEPO in this cell line were assessed by the percentage of viable cells, alterations of cell morphology, and the proliferative capability of the cells. In addition, we assessed the role of GEPO in lowering cellular oxidative stress and regulating expression of the anti-apoptotic protein Bcl2. The results showed rescue effects on the percentage of viable and proliferative cells among GEPO pretreated cells. Pretreatment with GEPO maintained the normal cell shape and ultrastructural morphology. Moreover, GEPO reduced the generation of reactive oxygen species in cells and activated expression of Bcl2, which are the major mechanisms in protection against cellular toxicity induced by AgNPs. In conclusion, our study showed that the cytotoxic effects from exposure to AgNPs can be prevented by GEPO. PMID:26929619

  18. Recombinant erythropoietin and blood transfusion in selected preterm infants

    PubMed Central

    Meyer, M; Sharma, E; Carsons, M

    2003-01-01

    Objectives: To comprehensively identify preterm infants likely to require blood transfusion and to investigate the effectiveness of recombinant erythropoietin in this high risk subgroup. Design: Double blind randomised controlled trial. Setting: Neonatal Intensive Care Unit, Middlemore Hospital, Auckland, New Zealand. Patients: Preterm infants < 33 weeks gestation and < 1700 g birth weight meeting specific criteria indicating a high possibility of requiring blood transfusion. Interventions: Predictors of blood transfusion were determined by analysis of preterm infants admitted to a neonatal intensive care unit over a two year period. Using the criteria developed, high risk infants entered the study and received erythropoietin or sham treatment until 34 weeks completed gestation. The sample size was calculated to detect a reduction of one blood transfusion per infant (significance level 5%, power 80%). Results: The selection criteria had a positive predictive value for transfusion of 91% and a negative predictive value of 94%. Mean birth weights and gestational ages were similar in the two groups. Absolute reticulocyte counts and haemoglobin values were higher in the group receiving erythropoietin. There was no significant difference in the number of blood transfusions received in the treatment and control groups. However, comparing transfusions given to < 1000 g infants after 30 days of age, there were significantly fewer transfusions in the erythropoietin group (mean (SD) 0.5 (0.7) in those receiving erythropoietin and 1.6 (1.1) in the controls). No adverse effects were noted. Conclusions: The selection criteria for the study were highly predictive of subsequent transfusion. In the group receiving erythropoietin, a reduction in transfusion requirements was apparent only in the < 1000 g birthweight group after 1 month of age. PMID:12496225

  19. Carbamylated Erythropoietin: A Prospective Drug Candidate for Neuroprotection.

    PubMed

    Chen, Jianmin; Yang, Zheng; Zhang, Xiao

    2015-01-01

    Carbamylated erythropoietin (cEpo), which is neuroprotective but lacks hematopoietic activity, has been attracting rising concerns. However, the cellular and molecular mechanisms involved in the process of neuroprotection of cEpo are not well known. Based on several recent reports, the neuroprotective effects of cEpo are illustrated, and signaling pathways involved in the different effects of erythropoietin and cEpo are discussed. These newly reported researches may shed new light on the development and application of cEpo, a prospective drug candidate for neuroprotection. PMID:26862298

  20. Carbamylated Erythropoietin: A Prospective Drug Candidate for Neuroprotection

    PubMed Central

    Chen, Jianmin; Yang, Zheng; Zhang, Xiao

    2015-01-01

    Carbamylated erythropoietin (cEpo), which is neuroprotective but lacks hematopoietic activity, has been attracting rising concerns. However, the cellular and molecular mechanisms involved in the process of neuroprotection of cEpo are not well known. Based on several recent reports, the neuroprotective effects of cEpo are illustrated, and signaling pathways involved in the different effects of erythropoietin and cEpo are discussed. These newly reported researches may shed new light on the development and application of cEpo, a prospective drug candidate for neuroprotection. PMID:26862298

  1. Expression of platelet-derived growth factor BB, erythropoietin and erythropoietin receptor in canine and feline osteosarcoma

    PubMed Central

    Meyer, F.R.L.; Steinborn, R.; Grausgruber, H.; Wolfesberger, B.; Walter, I.

    2015-01-01

    The discovery of expression of the erythropoietin receptor (EPO-R) on neoplastic cells has led to concerns about the safety of treating anaemic cancer patients with EPO. In addition to its endocrine function, the receptor may play a role in tumour progression through an autocrine mechanism. In this study, the expression of EPO, EPO-R and platelet-derived growth factor BB (PDGF-BB) was analysed in five feline and 13 canine osteosarcomas using immunohistochemistry (IHC) and reverse transcription polymerase chain reaction (RT-PCR). EPO expression was positive in all tumours by IHC, but EPO mRNA was only detected in 38% of the canine and 40% of the feline samples. EPO-R was expressed in all samples by quantitative RT-PCR (RT-qPCR) and IHC. EPO-R mRNA was expressed at higher levels in all feline tumours, tumour cell lines, and kidney when compared to canine tissues. PDGF-BB expression was variable by IHC, but mRNA was detected in all samples. To assess the functionality of the EPO-R on tumour cells, the proliferation of canine and feline osteosarcoma cell lines was evaluated after EPO administration using an alamarBlue assay and Ki67 immunostaining. All primary cell lines responded to EPO treatment in at least one of the performed assays, but the effect on proliferation was very low indicating only a weak responsiveness of EPO-R. In conclusion, since EPO and its receptor are expressed by canine and feline osteosarcomas, an autocrine or paracrine tumour progression mechanism cannot be excluded, although in vitro data suggest a minimal role of EPO-R in osteosarcoma cell proliferation. PMID:26189892

  2. Erythropoietin production by PDGFR-β(+) cells.

    PubMed

    Gerl, Katharina; Nolan, Karen A; Karger, Christian; Fuchs, Michaela; Wenger, Roland H; Stolt, Claus C; Willam, Carsten; Kurtz, Armin; Kurt, Birgül

    2016-08-01

    PDGFR-β-expressing cells of the kidneys are considered as a relevant site of erythropoietin (EPO) production. The origin of these cells, their contribution to renal EPO production, and if PDGFR-β-positive cells in other organs are also capable to express EPO are less clear. We addressed these questions in mice, in which hypoxia-inducible transcription factors were stabilized in PDGFR-β(+) cells by inducible deletion of the von Hippel-Lindau (Vhl) protein. Vhl deletion led to a 600-fold increase of plasma EPO concentration, 170-fold increase of renal EPO messenger RNA (mRNA) levels, and an increase of hematocrit values up to 70 %. Intrarenal localization of EPO-expressing cells coincided with the zonal heterogeneity and distribution of cells expressing PDGFR-β. Amongst a variety of extrarenal organs only adrenal glands showed significant EPO mRNA expression after Vhl deletion in PDGFR-β(+) cells. EPO mRNA, plasma EPO, and hematocrit fell to subnormal values if HIF-2α, but not HIF-1α, was deleted either alone or in combination with Vhl in PDGFR-β(+) cells. Treatment of mice with a prolyl-hydroxylase inhibitor caused an increase of EPO mRNA abundance and plasma EPO concentrations in wild-type mice and in mice lacking HIF-1α in PDGFR-β(+) cells but exerted no effect in mice lacking HIF-2α in PDGFR-β(+) cells. These findings suggest that PDGFR-β(+) cells are the only relevant site of EPO expression in the kidney and that HIF-2 is the essential transcription factor triggering EPO expression therein. Moreover, our findings suggest that PDGFR-β(+) cells elaborating EPO might arise from the metanephric mesenchyme, rather than from the neural crest. PMID:27220347

  3. Parsing Physiological Functions of Erythropoietin One Domain at a Time.

    PubMed

    Steinman, Lawrence

    2015-10-01

    A domain of erythropoietin (EPO), separate from the domain involved in red blood cell development, has been identified. This region of EPO has anti-inflammatory and neuroprotective effects. Use of a peptide sequence from this region provides the potential for an effective therapeutic without effects on erythropoiesis. PMID:26311151

  4. Erythropoietin, the biology of erythropoiesis and epoetin alfa. An overview.

    PubMed

    Bieber, E

    2001-05-01

    Erythropoietin, a glycoprotein hormone, is synthesized predominantly in the kidney and secreted by renal cortical interstitial cells in response to tissue hypoxia. Erythropoietin is the main regulator of the production of red blood cells. It functions in the recruitment and differentiation of erythroid progenitor cells and aids in their maintenance and survival. Erythropoietin also stimulates the synthesis of hemoglobin. In the last 15 years, the ready availability of recombinant human erythropoietin (r-HuEPO, epoetin alfa) has permitted the clinical investigation and application of this hormone to the treatment of anemia in various patient populations. Epoetin alfa has been shown to accelerate erythropoiesis and reduce allogeneic blood transfusion in major elective, noncardiac, nonvascular surgery and in certain anemic patients with chronic renal failure, nonmyeloid malignancies and human immunodeficiency virus infection. In addition to improving hematologic parameters, epoetin alfa therapy can enhance health-related quality of life in these patients. The success of epoetin alfa in treating anemia in other surgical populations suggests that it may be of benefit in treating the perioperative anemia that is highly prevalent in gynecologic surgery patients. Further investigation of the use of epoetin alfa in patients undergoing gynecologic surgery would increase awareness of its benefits for this patient population.

  5. [Ergometric results of r-erythropoietin treatment of hemodialysis patients].

    PubMed

    Hortian, B; Schmidt, R; Wüstenberg, P W; Dörp, E; Schumann, L; Winkler, R; Klinkmann, H

    1990-05-15

    To investigate the effect of partial correction of anemia in patients maintained by chronic intermittent hemodialysis on exercise capacity, patients underwent a bicycle acido-ergometry before and after treatment with recombinant human erythropoietin. The results demonstrate a (subjective) improvement of exercise capacity without any evidence for that in the pH values.

  6. The micronucleus test and erythropoiesis. Effects of erythropoietin and a mutagen on the ratio of polychromatic to normochromatic erythrocytes (P/N ratio).

    PubMed

    Suzuki, Y; Nagae, Y; Li, J; Sakaba, H; Mozawa, K; Takahashi, A; Shimizu, H

    1989-11-01

    It is considered that a decrease of the ratio of polychromatic erythrocytes (PCE) to normochromatic erythrocytes (NCE) (P/N) in the micronucleus test is an indicator of bone marrow toxicity induced by mutagens. However, the exact meaning of fluctuation in the P/N ratio is not yet known. We have studied this point by counting the total number of erythrocytes and nucleated cells in the bone marrow following various treatments. The P/N ratio decreased gradually with time after administration of mitomycin C. Our data suggest that the decrease of P/N ratio was attributable to an increase in the numbers of the denominator, i.e. NCE, caused by rapid differentiation and multiplication or denucleation of erythroblasts which remained in the bone marrow instead of entering the peripheral blood stream. A decrease of P/N ratio was also observed in the early phase after administration of erythropoietin, an agent which induces differentiation and multiplication of erythroblasts. This phenomenon might result from an increase of PCE delivery into the blood circulation. However, following the initial decrease, the P/N ratio increased gradually 48 h after administration of erythropoietin. It is supposed that this increase probably resulted from an increase in PCE in the bone marrow due to the direct effects of erythropoietin on erythropoiesis. The drastic change in erythropoiesis in the bone marrow induced by either mutagen or erythropoietin treatment will affect the fluctuations of the P/N ratio or the number of micronucleated erythrocytes per non-micronucleated erythocytes in the micronucleus test. This contrasts with the original explanation for such fluctuations which attributed them to replenishment of the marrow by peripheral blood. PMID:2516221

  7. Erythropoietin, uncertainty principle and cancer related anaemia

    PubMed Central

    Clark, Otavio; Adams, Jared R; Bennett, Charles L; Djulbegovic, Benjamin

    2002-01-01

    Background This study was designed to evaluate if erythropoietin (EPO) is effective in the treatment of cancer related anemia, and if its effect remains unchanged when data are analyzed according to various clinical and methodological characteristics of the studies. We also wanted to demonstrate that cumulative meta-analysis (CMA) can be used to resolve uncertainty regarding clinical questions. Methods Systematic Review (SR) of the published literature on the role of EPO in cancer-related anemia. A cumulative meta-analysis (CMA) using a conservative approach was performed to determine the point in time when uncertainty about the effect of EPO on transfusion-related outcomes could be considered resolved. Participants: Patients included in randomized studies that compared EPO versus no therapy or placebo. Main outcome measures: Number of patients requiring transfusions. Results Nineteen trials were included. The pooled results indicated a significant effect of EPO in reducing the number of patients requiring transfusions [odds ratio (OR) = 0.41; 95%CI: 0.33 to 0.5; p < 0.00001;relative risk (RR) = 0.61; 95% CI: 0.54 to 0.68]. The results remain unchanged after the sensitivity analyses were performed according to the various clinical and methodological characteristics of the studies. The heterogeneity was less pronounced when OR was used instead of RR as the measure of the summary point estimate. Analysis according to OR was not heterogeneous, but the pooled RR was highly heterogeneous. A stepwise metaregression analysis did point to the possibility that treatment effect could have been exaggerated by inadequacy in allocation concealment and that larger treatment effects are seen at hb level > 11.5 g/dl. We identified 1995 as the point in time when a statistically significant effect of EPO was demonstrated and after which we considered that uncertainty about EPO efficacy was resolved. Conclusion EPO is effective in the treatment of anemia in cancer patients. This

  8. Effect of Erythropoietin, Iron Deficiency and Iron Overload on Liver Matriptase-2 (TMPRSS6) Protein Content in Mice and Rats.

    PubMed

    Frýdlová, Jana; Přikryl, Petr; Truksa, Jaroslav; Falke, Lucas L; Du, Xin; Gurieva, Iuliia; Vokurka, Martin; Krijt, Jan

    2016-01-01

    Matriptase-2 (TMPRSS6) is an important negative regulator of hepcidin expression; however, the effects of iron overload or accelerated erythropoiesis on liver TMPRSS6 protein content in vivo are largely unknown. We determined TMPRSS6 protein content in plasma membrane-enriched fractions of liver homogenates by immunoblotting, using a commercial antibody raised against the catalytic domain of TMPRSS6. Plasma membrane-enriched fractions were obtained by centrifugation at 3000 g and washing. TMPRSS6 was detected in the 3000 g fraction as a 120 kDa full-length protein in both mice and rats. Feeding of iron-deficient diet as well as erythropoietin treatment increased TMPRSS6 protein content in rats and mice by a posttranscriptional mechanism; the increase in TMPRSS6 protein by erythropoietin was also observed in Bmp6-mutant mice. Administration of high doses of iron to mice (200, 350 and 700 mg/kg) decreased TMPRSS6 protein content. Hemojuvelin was detected in the plasma membrane-enriched fractions of control animals as a full length protein of approximately 52 kDa; in iron deficient animals, the full length protein was partially cleaved at the N-terminus, resulting in an additional weak band of approximately 47 kDa. In livers from hemojuvelin-mutant mice, TMPRSS6 protein content was strongly decreased, suggesting that intact hemojuvelin is necessary for stable TMPRSS6 expression in the membrane. Overall, the results demonstrate posttranscriptional regulation of liver TMPRSS6 protein by iron status and erythropoietin administration, and provide support for the interaction of TMPRSS6 and hemojuvelin proteins in vivo.

  9. Paraneoplastic Erythrocytosis of Colon Cancer, with Serum Erythropoietin within the Normal Reference Range

    PubMed Central

    Kitayama, Hiromitsu; Kondo, Tomohiro; Sugiyama, Junko; Hirayama, Michiaki; Oyamada, Yumiko; Tsuji, Yasushi

    2016-01-01

    Patient: Female, 75 Final Diagnosis: Erythropoietin-secreting colon cancer Symptoms: None Medication: — Clinical Procedure: Immunohistochemistry Specialty: Hematology Objective: Rare disease Background: Paraneoplastic erythrocytosis can be brought on by ectopic erythropoietin production usually in kidney, brain, and liver tumor with increase of serum erythropoietin level. We report here a paraneoplastic erythrocytosis of colon cancer with serum erythropoietin within the normal reference, which required an immunohistologic test for erythropoietin-antibody to be diagnosed. Case Report: Our case report was of a 75-year-old woman with erythrocytosis. Her hemoglobin and serum erythropoietin levels were 191 g/dL and 12.6 IU/L (reference range, 9.1–32.8), respectively. Colonoscopy revealed an advanced sigmoid colon tumor 20 mm in diameter. She underwent colectomy, and immunohistochemical examination showed the colon adenocarcinoma was focally positive for erythropoietin-antibody. One month after the surgery, her hemoglobin level decreased to 117 g/L. Conclusions: Colon cancer can cause paraneoplastic erythrocytosis, and it is important to consider not simply the absolute serum erythropoietin level but also the serum erythropoietin level relative to simultaneously measured hemoglobin level. We should include paraneoplastic erythrocytosis as a differential diagnosis in cases of high hemoglobin level unexplained by other diseases. PMID:27318703

  10. Survival and proliferative roles of erythropoietin beyond the erythroid lineage

    PubMed Central

    Noguchi, Constance Tom; Wang, Li; Rogers, Heather M.; Teng, Ruifeng; Jia, Yi

    2011-01-01

    Since the isolation and purification of erythropoietin (EPO) in 1977, the essential role of EPO for mature red blood cell production has been well established. The cloning and production of recombinant human EPO led to its widespread use in treating patients with anaemia. However, the biological activity of EPO is not restricted to regulation of erythropoiesis. EPO receptor (EPOR) expression is also found in endothelial, brain, cardiovascular and other tissues, although at levels considerably lower than that of erythroid progenitor cells. This review discusses the survival and proliferative activity of EPO that extends beyond erythroid progenitor cells. Loss of EpoR expression in mouse models provides evidence for the role of endogenous EPO signalling in nonhaematopoietic tissue during development or for tissue maintenance and/or repair. Determining the extent and distribution of receptor expression provides insights into the potential protective activity of erythropoietin in brain, heart and other nonhaematopoietic tissues. PMID:19040789

  11. Mesenchymal Stromal Cells Engineered to Express Erythropoietin Induce Anti-erythropoietin Antibodies and Anemia in Allorecipients

    PubMed Central

    Campeau, Philippe M; Rafei, Moutih; François, Moïra; Birman, Elena; Forner, Kathy-Ann; Galipeau, Jacques

    2008-01-01

    Autologous bone marrow mesenchymal stromal cells (MSCs) have been successfully used for the delivery of erythropoietin (EPO) in murine models of anemia and myocardial infarction. For clinical applications where a transient effect would be adequate, such as myocardial infarction, the use of EPO-engineered universal donor allogeneic MSCs would be a substantial convenience. We thus investigated whether MSCs from C57BL/6 mice would permit robust transient EPO delivery in normal BALB/c allorecipients. Implantation of MSCs overexpressing murine EPO led to increases in hematocrit in syngeneic and allogeneic mice, but the latter eventually developed severe anemia due to acquired neutralizing anti-EPO antibodies. As MSCs constitutively produce the CCL2 chemokine which may behave as an adjuvant to the anti-EPO immune response, experiments were performed using EPO-engineered MSCs derived from CCL2–/– mice and similar results were obtained. In conclusion, MHC-mismatched MSCs can break the tolerance to autoantigens and lead to the development of pathogenic autoantibodies. PMID:19088705

  12. Erythropoietin and the use of a transgenic model of erythropoietin-deficient mice

    PubMed Central

    Pichon, Aurélien; Jeton, Florine; El Hasnaoui-Saadani, Raja; Hagström, Luciana; Launay, Thierry; Beaudry, Michèle; Marchant, Dominique; Quidu, Patricia; Macarlupu, Jose-Luis; Favret, Fabrice; Richalet, Jean-Paul; Voituron, Nicolas

    2016-01-01

    Despite its well-known role in red blood cell production, it is now accepted that erythropoietin (Epo) has other physiological functions. Epo and its receptors are expressed in many tissues, such as the brain and heart. The presence of Epo/Epo receptors in these organs suggests other roles than those usually assigned to this protein. Thus, the aim of this review is to describe the effects of Epo deficiency on adaptation to normoxic and hypoxic environments and to suggest a key role of Epo on main physiological adaptive functions. Our original model of Epo-deficient (Epo-TAgh) mice allowed us to improve our knowledge of the possible role of Epo in O2 homeostasis. The use of anemic transgenic mice revealed Epo as a crucial component of adaptation to hypoxia. Epo-TAgh mice survive well in hypoxic conditions despite low hematocrit. Furthermore, Epo plays a key role in neural control of ventilatory acclimatization and response to hypoxia, in deformability of red blood cells, in cerebral and cardiac angiogenesis, and in neuro- and cardioprotection. PMID:27800506

  13. Collaborative study for the establishment of erythropoietin BRP batch 4.

    PubMed

    Burns, C; Bristow, A F; Daas, A; Costanzo, A

    2015-01-01

    The European Pharmacopoeia (Ph. Eur.) Biological Reference Preparation (BRP) for erythropoietin (EPO) is used as a working standard for potency determination of EPO preparations by in vivo bioassay as prescribed in the Ph. Eur. monograph Erythropoietin concentrated solution (1316). The BRP batch 3 was calibrated in 2006 and its stocks are depleted. The European Directorate for the Quality of Medicines & HealthCare (EDQM) thus initiated a project to calibrate a replacement batch in International Units against the WHO 3(rd) International Standard (IS) for Erythropoietin, recombinant, for bioassay (11/170). A Ph. Eur. Chemical Reference Substance (CRS) was established recently for use as reference in some of the physicochemical tests prescribed in the monograph. Therefore, the EPO BRP batch 4 was only calibrated for the normocythaemic and polycythaemic mouse in vivo bioassays described in the Assay section of the Ph. Eur. monograph (1316). The collaborative study involved seven laboratories from Europe, the USA and South America. The results confirmed that the candidate BRP (cBRP) is suitable for use as a reference preparation in the potency determination of EPO medicinal products by bioassay (using the normocythaemic or polycythaemic mouse methods). The outcome of the study enabled the Ph. Eur. Commission to establish the proposed standard as erythropoietin BRP batch 4 in November 2014 for use as a reference preparation solely for the polycythaemic and normocythaemic mouse bioassay, with an assigned potency of 13 000 IU/vial. Furthermore, the potency of BRP3 was confirmed during the study, thus warranting a good continuity of the IU.

  14. Use of erythropoietin in emergencies: massive intoxication by chloramines.

    PubMed

    Lorenzo, I; Medina, M; Calderon, P; Castro, S; Lazaro, R

    1996-01-01

    Recombinant human Erythropoietin (rHuEPO) is normally used to correct anaemia in patients with End Stage Renal Disease (ESRD), that are in Regular Dialysis Treatment (RDT). This anaemia is usually due to the existence of two factors: A decrease in the erythropoiesis of the bone marrow and an increase in peripheral haemolysis and, consequently, a decrease in the life span of the red cells. PMID:10723309

  15. Acute Vhl gene inactivation induces cardiac HIF-dependent erythropoietin gene expression.

    PubMed

    Miró-Murillo, Marta; Elorza, Ainara; Soro-Arnáiz, Inés; Albacete-Albacete, Lucas; Ordoñez, Angel; Balsa, Eduardo; Vara-Vega, Alicia; Vázquez, Silvia; Fuertes, Esther; Fernández-Criado, Carmen; Landázuri, Manuel O; Aragonés, Julián

    2011-01-01

    Von Hippel Lindau (Vhl) gene inactivation results in embryonic lethality. The consequences of its inactivation in adult mice, and of the ensuing activation of the hypoxia-inducible factors (HIFs), have been explored mainly in a tissue-specific manner. This mid-gestation lethality can be also circumvented by using a floxed Vhl allele in combination with an ubiquitous tamoxifen-inducible recombinase Cre-ER(T2). Here, we characterize a widespread reduction in Vhl gene expression in Vhl(floxed)-UBC-Cre-ER(T2) adult mice after dietary tamoxifen administration, a convenient route of administration that has yet to be fully characterized for global gene inactivation. Vhl gene inactivation rapidly resulted in a marked splenomegaly and skin erythema, accompanied by renal and hepatic induction of the erythropoietin (Epo) gene, indicative of the in vivo activation of the oxygen sensing HIF pathway. We show that acute Vhl gene inactivation also induced Epo gene expression in the heart, revealing cardiac tissue to be an extra-renal source of EPO. Indeed, primary cardiomyocytes and HL-1 cardiac cells both induce Epo gene expression when exposed to low O(2) tension in a HIF-dependent manner. Thus, as well as demonstrating the potential of dietary tamoxifen administration for gene inactivation studies in UBC-Cre-ER(T2) mouse lines, this data provides evidence of a cardiac oxygen-sensing VHL/HIF/EPO pathway in adult mice.

  16. [Study of the mechanism of erythropoietin effect on energy metabolism in the bone marrow].

    PubMed

    Zinov'ev, Iu V

    1976-01-01

    In extracts of rabbit bone marrow cells was studied effect of erythropoietine on the activity of some enzymes (hexokinase, phosphoglucomutase, phosphohexoisomerase, lactate dehydrogenase, glucoso-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase and NADP-reductase). The NADP-reductase activity was increased under the effect of erythropoietine; the activities of other enzymes studied was not altered.

  17. Phagocyte respiratory burst activates macrophage erythropoietin signalling to promote acute inflammation resolution

    PubMed Central

    Luo, Bangwei; Wang, Jinsong; Liu, Zongwei; Shen, Zigang; Shi, Rongchen; Liu, Yu-Qi; Liu, Yu; Jiang, Man; Wu, Yuzhang; Zhang, Zhiren

    2016-01-01

    Inflammation resolution is an active process, the failure of which causes uncontrolled inflammation which underlies many chronic diseases. Therefore, endogenous pathways that regulate inflammation resolution are fundamental and of wide interest. Here, we demonstrate that phagocyte respiratory burst-induced hypoxia activates macrophage erythropoietin signalling to promote acute inflammation resolution. This signalling is activated following acute but not chronic inflammation. Pharmacological or genetical inhibition of the respiratory burst suppresses hypoxia and macrophage erythropoietin signalling. Macrophage-specific erythropoietin receptor-deficient mice and chronic granulomatous disease (CGD) mice, which lack the capacity for respiratory burst, display impaired inflammation resolution, and exogenous erythropoietin enhances this resolution in WT and CGD mice. Mechanistically, erythropoietin increases macrophage engulfment of apoptotic neutrophils via PPARγ, promotes macrophage removal of debris and enhances macrophage migration to draining lymph nodes. Together, our results provide evidences of an endogenous pathway that regulates inflammation resolution, with important implications for treating inflammatory conditions. PMID:27397585

  18. Effects Of Recombinant Human Erythropoietin On Platelet Activation In Acute Myocardial Infarction: Results Of A Double-Blind, Placebo-Controlled Randomized Trial

    PubMed Central

    Tang, Yi-Da; Hasan, Faisal; Giordano, Frank J.; Pfau, Stephen; Rinder, Henry M.; Katz, Stuart D.

    2009-01-01

    Background Erythropoietin mitigates myocardial damage and improves ventricular performance after experimental ischemic injury. This study assessed safety and efficacy markers relevant to the biological activity of recombinant human erythropoietin (rHuEpo) in patients with acute myocardial infarction. Methods We conducted a prospective, placebo-controlled, randomized, double-blind trial to determine the effects of intravenous rHuEpo (200 U/kg daily for 3 consecutive days) on measures of platelet and endothelial cell activation, soluble Fas ligand, and peripheral blood mononuclear cell (PBMC) expression of angiogenesis signaling proteins in 44 subjects with acute myocardial infarction (MI) treated with aspirin and clopidogrel after successful percutaneous coronary intervention. Results rHuEpo did not alter bleeding time, platelet function assay closure time, von Willebrand factor levels, soluble P-selectin, or soluble Fas ligand levels when compared with placebo. By contrast, rHuEpo significantly increased expression of erythropoietin receptor, vascular endothelial growth factor receptor Flt-1, and phosphorylated phosphatidylinositol 3-kinase in PBMC’s when compared with placebo (all p<0.05). Conclusions In acute MI patients treated with aspirin and clopidogrel, short-term administration of rHuEpo did not alter markers of platelet and endothelial cell activation associated with thrombosis, yet did increase expression of angiogenesis signaling proteins in PBMC’s when compared with placebo. These data provide preliminary evidence of safety and biologic activity of rHuEpo at this dosing and support continued enrollment in ongoing efficacy trials. PMID:19958860

  19. AMPK is involved in mediation of erythropoietin influence on metabolic activity and reactive oxygen species production in white adipocytes.

    PubMed

    Wang, Li; Di, Lijun; Noguchi, Constance Tom

    2014-09-01

    Erythropoietin, discovered for its indispensable role during erythropoiesis, has been used in therapy for selected red blood cell disorders in erythropoietin-deficient patients. The biological activities of erythropoietin have been found in animal models to extend to non-erythroid tissues due to the expression of erythropoietin receptor. We previously demonstrated that erythropoietin promotes metabolic activity and white adipocytes browning to increase mitochondrial function and energy expenditure via peroxisome proliferator-activated receptor alpha and Sirtuin1. Here we report that AMP-activated protein kinase was activated by erythropoietin possibly via Ca(2+)/calmodulin-dependent protein kinase kinase in adipocytes as well as in white adipose tissue from diet induced obese mice. Erythropoietin increased cellular nicotinamide adenine dinucleotide via increased AMP-activated protein kinase activity, possibly leading to Sirtuin1 activation. AMP-activated protein kinase knock down reduced erythropoietin mediated increase in cellular oxidative function including the increased oxygen consumption rate, fatty acid utilization and induction of key metabolic genes. Under hypoxia, adipocytes were found to generate more reactive oxygen species, and erythropoietin reduced the reactive oxygen species and increased antioxidant gene expression, suggesting that erythropoietin may provide protection from oxidative stress in adipocytes. Erythropoietin also reversed increased nicotinamide adenine dinucleotide by hypoxia via increased AMP-activated protein kinase. Additionally, AMP-activated protein kinase is found to be involved in erythropoietin stimulated increase in oxygen consumption rate, fatty acid oxidation and mitochondrial gene expression. AMP-activated protein kinase knock down impaired erythropoietin stimulated increases in antioxidant gene expression. Collectively, our findings identify the AMP-activated protein kinase involvement in erythropoietin signaling in

  20. Nonclinical evaluation of the potential for mast cell activation by an erythropoietin analog.

    PubMed

    Weaver, James L; Boyne, Michael; Pang, Eric; Chimalakonda, Krishna; Howard, Kristina E

    2015-09-15

    The erythropoietin analog peginesatide was withdrawn from marketing due to unexpected severe anaphylactic reactions associated with administration of the multi-use formulation. The adverse events occurred rapidly following the first ever administration of the drug with most affected patients becoming symptomatic in less than 30min. This is most consistent with an anaphylactoid reaction due to direct activation of mast cells. Laboratory evaluation was undertaken using rat peritoneal mast cells as the model system. Initial studies showed that high concentrations of the formulated drug as well as formulated vehicle alone could cause mast cell degranulation as measured by histamine release. The purified active drug was not able to cause histamine release whereas the vehicle filtrate and lab created drug vehicle were equally potent at causing histamine release. Individual formulations of vehicle leaving one component out showed that histamine release was due to phenol. Dose response studies with phenol showed a very sharp dose response curve that was similar in three buffer systems. Cellular analysis by flow cytometry showed that the histamine release was not due to cell death, and that changes in light scatter parameters consistent with degranulation were rapidly observed. Limited testing with primary human mast cells showed a similar dose response of histamine release with exposure to phenol. To provide in vivo confirmation, rats were injected with vehicle formulated with various concentrations of phenol via a jugular vein cannula. Significant release of histamine was detected in blood samples taken 2min after dosing at the highest concentrations tested. PMID:26079829

  1. Anemia in the preterm infant: Erythropoietin versus erythrocyte transfusion — It’s not that simple

    PubMed Central

    Von Kohorn, Isabelle; Ehrenkranz, Richard A.

    2009-01-01

    SYNOPSIS Since the late 1980s recombinant human erythropoietin (r-Epo) has been studied as an alternative to packed red blood cell (RBC) transfusion for the treatment of anemia of prematurity in very low birth weight (VLBW, <1500 grams) infants. Initial trials and reports focused on r-Epo’s ability to prevent or treat anemia of prematurity with the goal of eliminating RBC transfusion, but achieved limited success. Reduced volumes of blood sampling for laboratory tests and improved blood banking techniques have decreased the need for RBC transfusion. New concerns about the safety of r-Epo administration have emerged. Past cost-benefit analyses of r-Epo administration versus transfusion for the treatment of anemia of prematurity have been nearly balanced. Autologous transfusion, blood-sparing technologies, changes in RBC transfusion technique and safety, and further elucidation of the risk-benefit ratio of r-Epo therapy may change the cost-benefit analysis. The jury is still out with regard to the role of r-Epo therapy in the VLBW population. PMID:19161869

  2. Combination of exercise training and erythropoietin prevents cancer-induced muscle alterations.

    PubMed

    Pin, Fabrizio; Busquets, Silvia; Toledo, Miriam; Camperi, Andrea; Lopez-Soriano, Francisco J; Costelli, Paola; Argilés, Josep M; Penna, Fabio

    2015-12-22

    Cancer cachexia is a syndrome characterized by loss of skeletal muscle mass, inflammation, anorexia and anemia, contributing to patient fatigue and reduced quality of life. In addition to nutritional approaches, exercise training (EX) has been proposed as a suitable tool to manage cachexia. In the present work the effect of mild exercise training, coupled to erythropoietin (EPO) administration to prevent anemia, has been tested in tumor-bearing mice. In the C26 hosts, acute exercise does not prevent and even worsens muscle wasting. Such pattern is prevented by EPO co-administration or by the adoption of a chronic exercise protocol. EX and EPO co-treatment spares oxidative myofibers from atrophy and counteracts the oxidative to glycolytic shift, inducing PGC-1α. LLC hosts are responsive to exercise and their treatment with the EX-EPO combination prevents the loss of muscle strength and the onset of mitochondrial ultrastructural alterations, while increases muscle oxidative capacity and intracellular ATP content, likely depending on PGC-1α induction and mitophagy promotion. Consistently, muscle-specific PGC-1α overexpression prevents LLC-induced muscle atrophy and Atrogin-1 hyperexpression. Overall, the present data suggest that low intensisty exercise can be an effective tool to be included in combined therapeutic approaches against cancer cachexia, provided that anemia is coincidently treated in order to enhance the beneficial action of exercise.

  3. Biological evaluation of recombinant human erythropoietin in pharmaceutical products.

    PubMed

    Ramos, A S; Schmidt, C A; Andrade, S S; Fronza, M; Rafferty, B; Dalmora, S L

    2003-11-01

    The potencies of mammalian cell-derived recombinant human erythropoietin pharmaceutical preparations, from a total of five manufacturers, were assessed by in vivo bioassay using standardized protocols. Eight-week-old normocythemic mice received a single subcutaneous injection followed by blood sampling 96 h later or multiple daily injections with blood sampling 24 h after the last injection. Reticulocyte counting by microscopic examination was employed as the end-point using the brilliant cresyl blue or selective hemolysis methods, together with automated flow cytometry. Different injection schedules were investigated and dose-response curves for the European Pharmacopoeia Biological Reference Preparation of erythropoietin were compared. Manual and automated methods of reticulocyte counting were correlated with respect to assay validity and precision. Using 8 mice per treatment group, intra-assay precision determined for all of the assays in the study showed coefficients of variation of 12.1-28.4% for the brilliant cresyl blue method, 14.1-30.8% for the selective hemolysis method and 8.5-19.7% for the flow cytometry method. Applying the single injection protocol, a combination of at least two independent assays was required to achieve the precision potency and confidence limits indicated by the manufacturers, while the multiple daily injection protocol yielded the same acceptable results within a single assay. Although the latter protocol using flow cytometry for reticulocyte counting gave more precise and reproducible results (intra-assay coefficients of variation: 5.9-14.2%), the well-characterized manual methods provide equally valid alternatives for the quality control of recombinant human erythropoietin therapeutic products.

  4. Effects of Intraosseous Erythropoietin during Hemorrhagic Shock in Swine

    PubMed Central

    Borovnik-Lesjak, Vesna; Whitehouse, Kasen; Baetiong, Alvin; Miao, Yang; Currie, Brian M.; Velmurugan, Sathya; Radhakrishnan, Jeejabai; Gazmuri, Raúl J.

    2014-01-01

    Objective To determine whether erythropoietin given during hemorrhagic shock (HS) ameliorates organ injury while improving resuscitation and survival. Methods Three series of 24 pigs each were studied. In an initial series, 50% of the blood volume (BV) was removed in 30 minutes and normal saline (threefold the blood removed) started at minute 90 infusing each third in 30, 60, and 150 minutes with shed blood reinfused at minute 330 (HS-50BV). In a second series, the same HS-50BV protocol was used but removing an additional 15% of BV from minute 30 to 60 (HS-65BV). In a final series, blood was removed as in HS-65BV and intraosseous vasopressin given from minute 30 (0.04 U/kg min−1) until start of shed blood reinfusion at minute 150 (HS-65BV+VP). Normal saline was reduced to half the blood removed and given from minute 90 to 120 in half of the animals. In each series, animals were randomized 1∶1 to receive erythropoietin (1,200 U/kg) or control solution intraosseously after removing 10% of the BV. Results In HS-50BV, O2 consumption remained near baseline yielding minimal lactate increases, 88% resuscitability, and 60% survival at 72 hours. In HS-65BV, O2 consumption was reduced and lactate increased yielding 25% resuscitability. In HS-65BV+VP, vasopressin promoted hemodynamic stability yielding 92% resuscitability and 83% survival at 72 hours. Erythropoietin did not affect resuscitability or subsequent survival in any of the series but increased interleukin-10, attenuated lactate increases, and ameliorated organ injury based on lesser troponin I, AST, and ALT increases and lesser neurological deficits in the HS-65BV+VP series. Conclusions Erythropoietin given during HS in swine failed to alter resuscitability and 72 hour survival regardless of HS severity and concomitant treatment with fluids and vasopressin but attenuated acute organ injury. The studies also showed the efficacy of vasopressin and restrictive fluid resuscitation for hemodynamic stabilization and

  5. On the in vivo action of erythropoietin: a quantitative analysis.

    PubMed

    Papayannopoulou, T; Finch, C A

    1972-05-01

    The composite response of the erythron to exogenous erythropoietin has been studied in normal, splenectomized, and polycythemic mice. After stimulation the normal animal doubled its marrow nucleated red cells by the 3rd day with little further change by the 5th. Nucleated red cells within the spleen began to increase sharply on the 2nd day and, by the 5th, exceeded those in the marrow. The total nucleated erythroid response represented a fourfold increase. Reticulocytes lagged behind the expansion of the nucleated red cell mass, but by the 5th day the original ratio was re-established. Hemoglobin synthesis was increased, but the ratio of hemoglobin synthesized in nucleated red cells and reticulocytes was basically unchanged. Early displacement of marrow reticulocytes into circulation and the production of a larger red cell also occurred. No evidence of a change in the number of erythroid mitoses was found; only a slight decrease in the average cell cycle time was demonstrated. Thus, whereas erythropoietin stimulation induced several changes in erythropoiesis, the increased number of cells entering into the maturing pool appeared to be of greatest quantitative significance.Splenectomy reduced the proliferative response of the erythron over 5 days stimulation to three-fourths that found in the normal animal. This difference, also reflected in a proportionately lower reticulocyte response and increment in circulating red cell mass, suggests that erythropoiesis within the mouse marrow is spatially or otherwise restricted and that the spleen provided a supplemental area of erythroid expansion.

  6. On the in vivo action of erythropoietin: a quantitative analysis

    PubMed Central

    Papayannopoulou, Thalia; Finch, Clement A.

    1972-01-01

    The composite response of the erythron to exogenous erythropoietin has been studied in normal, splenectomized, and polycythemic mice. After stimulation the normal animal doubled its marrow nucleated red cells by the 3rd day with little further change by the 5th. Nucleated red cells within the spleen began to increase sharply on the 2nd day and, by the 5th, exceeded those in the marrow. The total nucleated erythroid response represented a fourfold increase. Reticulocytes lagged behind the expansion of the nucleated red cell mass, but by the 5th day the original ratio was re-established. Hemoglobin synthesis was increased, but the ratio of hemoglobin synthesized in nucleated red cells and reticulocytes was basically unchanged. Early displacement of marrow reticulocytes into circulation and the production of a larger red cell also occurred. No evidence of a change in the number of erythroid mitoses was found; only a slight decrease in the average cell cycle time was demonstrated. Thus, whereas erythropoietin stimulation induced several changes in erythropoiesis, the increased number of cells entering into the maturing pool appeared to be of greatest quantitative significance. Splenectomy reduced the proliferative response of the erythron over 5 days stimulation to three-fourths that found in the normal animal. This difference, also reflected in a proportionately lower reticulocyte response and increment in circulating red cell mass, suggests that erythropoiesis within the mouse marrow is spatially or otherwise restricted and that the spleen provided a supplemental area of erythroid expansion. PMID:5020431

  7. Erythropoietin Activates Mitochondrial Biogenesis and Couples Red Cell Mass to Mitochondrial Mass in the Heart

    EPA Science Inventory

    RATIONALE: Erythropoietin (EPO) is often administered to cardiac patients with anemia, particularly from chronic kidney disease, and stimulation of erythropoiesis may stabilize left ventricular and renal function by recruiting protective effects beyond the correction of anemia. O...

  8. Comparison of Hemoglobin Levels Before and After Hemodialysis and Their Effects on Erythropoietin Dosing and Cost

    PubMed Central

    Sagheb, Mohammad Mahdi; Fallahzadeh, Mohammad Amin; Moaref, Alireza; Fallahzadeh, Mohammad Hossein; Dormanesh, Banafshe

    2016-01-01

    Background Hemoglobin levels measured after hemodialysis, as compared to hemoglobin levels measured before hemodialysis, are suggested to be a more accurate reflection of the hemoglobin levels between hemodialysis sessions, and to be a better reference point for adjusting erythropoietin dosing. Objectives The aim of this study was to compare the hemoglobin levels before and after hemodialysis, to calculate the required erythropoietin doses based on these levels, and to develop a model to predict effective erythropoietin dosing. Patients and Methods In this cross-sectional study, the hemoglobin levels of 52 patients with end-stage renal disease were measured before and after hemodialysis. The required erythropoietin doses and the differences in cost were calculated based on the hemoglobin levels before and after hemodialysis. A model to predict the adjusted erythropoietin dosages based on post-hemodialysis hemoglobin levels was proposed. Results Hemoglobin levels measured after hemodialysis were significantly higher than the hemoglobin levels before hemodialysis (11.1 ± 1.1 vs. 11.9 ± 1.2 g/dL, P < 0.001, 7% increase). The mean required erythropoietin dose based on post-hemodialysis hemoglobin levels was significantly lower than the corresponding erythropoietin dose based on pre-hemodialysis hemoglobin levels (10947 ± 6820 vs. 12047 ± 7542 U/week, P < 0.001, 9% decrease). The cost of erythropoietin was also significantly lower when post-hemodialysis levels were used (15.96 ± 9.85 vs. 17.57 ± 11.00 dollars/patient/week, P < 0.001). This translated into 83.72 dollars/patient/year in cost reduction. The developed model for predicting the required dosage is: Erythropoietin (U/week) = 43540.8 + (-2734.8) × Post-hemodialysis Hb* (g/dL). [(R2) = 0.221; *P < 0.001]. Conclusions Using post-hemodialysis hemoglobin levels as a reference point for erythropoietin dosing can result in significant dose and cost reduction, and can protect hemodialysis patients from

  9. Erythropoietin reduces storage lesions and decreases apoptosis indices in blood bank red blood cells

    PubMed Central

    Penuela, Oscar Andrés; Palomino, Fernando; Gómez, Lina Andrea

    2015-01-01

    Background Recent evidence shows a selective destruction of the youngest circulating red blood cells (neocytolysis) trigged by a drop in erythropoietin levels. Objective The aim of this study was to evaluate the effect of recombinant human erythropoietin beta on the red blood cell storage lesion and apoptosis indices under blood bank conditions. Methods Each one of ten red blood cell units preserved in additive solution 5 was divided in two volumes of 100 mL and assigned to one of two groups: erythropoietin (addition of 665 IU of recombinant human erythropoietin) and control (isotonic buffer solution was added). The pharmacokinetic parameters of erythropoietin were estimated and the following parameters were measured weekly, for six weeks: Immunoreactive erythropoietin, hemolysis, percentage of non-discocytes, adenosine triphosphate, glucose, lactate, lactate dehydrogenase, and annexin-V/esterase activity. The t-test or Wilcoxon's test was used for statistical analysis with significance being set for a p-value <0.05. Results Erythropoietin, when added to red blood cell units, has a half-life >6 weeks under blood bank conditions, with persistent supernatant concentrations of erythropoietin during the entire storage period. Adenosine triphosphate was higher in the Erythropoietin Group in Week 6 (4.19 ± 0.05 μmol/L vs. 3.53 ± 0.02 μmol/L; p-value = 0.009). The number of viable cells in the Erythropoietin Group was higher than in the Control Group (77% ± 3.8% vs. 71% ± 2.3%; p-value <0.05), while the number of apoptotic cells was lower (9.4% ± 0.3% vs. 22% ± 0.8%; p-value <0.05). Conclusions Under standard blood bank conditions, an important proportion of red blood cells satisfy the criteria of apoptosis. Recombinant human erythropoietin beta seems to improve storage lesion parameters and mitigate apoptosis. PMID:26969770

  10. Nitric oxide prevents cardiovascular disease and determines survival in polyglobulic mice overexpressing erythropoietin.

    PubMed

    Ruschitzka, F T; Wenger, R H; Stallmach, T; Quaschning, T; de Wit, C; Wagner, K; Labugger, R; Kelm, M; Noll, G; Rülicke, T; Shaw, S; Lindberg, R L; Rodenwaldt, B; Lutz, H; Bauer, C; Lüscher, T F; Gassmann, M

    2000-10-10

    Nitric oxide (NO) induces vasodilatatory, antiaggregatory, and antiproliferative effects in vitro. To delineate potential beneficial effects of NO in preventing vascular disease in vivo, we generated transgenic mice overexpressing human erythropoietin. These animals induce polyglobulia known to be associated with a high incidence of vascular disease. Despite hematocrit levels of 80%, adult transgenic mice did not develop hypertension or thromboembolism. Endothelial NO synthase levels, NO-mediated endothelium-dependent relaxation and circulating and vascular tissue NO levels were markedly increased. Administration of the NO synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) led to vasoconstriction of peripheral resistance vessels, hypertension, and death of transgenic mice, whereas wild-type siblings developed hypertension but did not show increased mortality. L-NAME-treated polyglobulic mice revealed acute left ventricular dilatation and vascular engorgement associated with pulmonary congestion and hemorrhage. In conclusion, we here unequivocally demonstrate that endothelial NO maintains normotension, prevents cardiovascular dysfunction, and critically determines survival in vivo under conditions of increased hematocrit.

  11. Nonerythropoietic Erythropoietin-Derived Peptide Suppresses Adipogenesis, Inflammation, Obesity and Insulin Resistance.

    PubMed

    Liu, Yuqi; Luo, Bangwei; Shi, Rongchen; Wang, Jinsong; Liu, Zongwei; Liu, Wei; Wang, Shufeng; Zhang, Zhiren

    2015-10-13

    Erythropoietin (EPO) has been identified as being crucial for obesity modulation; however, its erythropoietic activity may limit its clinical application. EPO-derived Helix B-surface peptide (pHBSP) is nonerythrogenic but has been reported to retain other functions of EPO. The current study aimed to evaluate the effects and potential mechanisms of pHBSP in obesity modulation. We found that pHBSP suppressed adipogenesis, adipokine expression and peroxisome proliferator-activated receptor γ (PPARγ) levels during 3T3-L1 preadipocyte maturation through the EPO receptor (EPOR). In addition, also through EPOR, pHBSP attenuated macrophage inflammatory activation and promoted PPARγ expression. Furthermore, PPARγ deficiency partly ablated the anti-inflammatory activity of pHBSP in macrophages. Correspondingly, pHBSP administration to high-fat diet (HFD)-fed mice significantly improved obesity, insulin resistance (IR) and adipose tissue inflammation without stimulating hematopoiesis. Therefore, pHBSP can significantly protect against obesity and IR partly by inhibiting adipogenesis and inflammation. These findings have therapeutic implications for metabolic disorders, such as obesity and diabetes.

  12. Particle-induced erythropoietin-independent effects of erythroid precursor cells in murine bone marrow.

    PubMed

    Ploemacher, R E; van Soest, P L; Wagemaker, G; van 't Hull, E

    1979-09-01

    A possible regulatory action of phagocytic cells on erythropoiesis was investigated by infusion of inert polystyrene latex particles (LAT). LAT appeared to induce changes in the femoral content of erythroid progenitor cells. These changes were most pronounced in primitive erythroid progenitor cells (BFUe) and appeared to be gradually damped in more differentiated populations (CFUe and erythroblasts). LAT did not influence granulocyte/macrophage progenitor cells (CFUc). The effects of LAT could not be attributed to changes in the systemic erythropoietin (EP) concentration. Administration of dexamethason nullified the effect of low doses of LAT, suggesting that phagocytosis of the particles is essential to the observed effects. Erythroid burst formation was previously found to be dependent on a bone marrow associated activity, termed BFA (burst feeder activity). BFA acts as an in vitro inducer of EP-responsiveness in BFUe. In this study it was found that LAT-induced changes in femoral erythroid progenitor cell content were characteristically preceded by corresponding changes in BFA. It was concluded that BFA-associated cells probably play a role in vivo in the early differentiation of erythroid progenitor cells. The present data are interpreted as direct in vivo evidence supporting a two-step regulatory model operating in erythropoiesis and provide evidence that phagocytic cells are a component of the erythroid haemopoietic inductive micro-environment. PMID:519701

  13. Erythropoietin-independent regeneration of erythroid progenitor cells following multiple injections of hydroxyurea.

    PubMed

    Wagemaker, G; Visser, T P

    1980-09-01

    It wa shown previously that colony formation in vitro by early erythroid progenitor cells (BFUe) requires sequential stimulation with a specific glycoprotein termed BFA and erythropoietin (EP). The action exerted by BFA was characterized as induction of proliferation in BFUe resulting after several cell divisions in EP-responsive progeny. The present study is directed at detection of EP-independent regulation of erythroid progenitor cells in vivo. Haemopoietic regeneration was induced by multiple administrations of hydroxyurea (HU). The femoral regeneration patterns of haemopoietic stem cells (CFUs), granulocyte/macrophage progenitor cells (CFUgm) and erythroid progenitor cells (BFUe, day 3 BFUe and CFUe) were studied in hypertransfused mice in comparison to nontransfused controls. The results show that (1) the phase of exponential regeneration of none of the cell populations studied is affected by hypertransfusion; (2) each of these cell populations exhibit a distinct regeneration pattern, indicating that they behave as separate functional entities; and (3) the three erythroid cell populations are suppressed by hypertransfusion in the post-exponential phase of regeneration in contrast to CFUs and CFUgm. The results support a two-regulator model of erythropoiesis. PMID:7459981

  14. Reduced serum hydroxyl radical scavenging activity in erythropoietin therapy resistant renal anemia.

    PubMed

    Hirayama, Aki; Nagase, Sohji; Gotoh, Michihiro; Ueda, Atsushi; Ishizu, Takashi; Yoh, Keigyou; Aoyagi, Kazumasa; Terao, Junji; Koyama, Akio

    2002-11-01

    Relation between anemia resistant to recombinant human erythropoietin (rHuEPO) therapy and the oxidative stress in hemodialysis (HD) patients was investigated. Stable HD patients who had consistent hemoglobin concentrations on a constant dose of rHuEPO were studied. Patients were excluded if there were factors that might affect hemopoiesis or administration of antioxidant supplements. Patients were classified into three groups: High (9000 U/week), Low (1500-4500 U/week) and No rHuEPO group. Thiobarbituric acid reactive substances (TBARS) of sera and erythrocyte were examined. Serum superoxide and hydroxyl radical scavenging activities were measured using electron spin resonance. TBARS in the erythrocyte was higher in High rHuEPO group compared with No rHuEPO group, though the serum TBARS were similar. A diminution of serum hydroxyl radical scavenging activity was observed in High rHuEPO group. Hydroxyl radical signal intensity showed a strong correlation with the serum ferritin in High rHuEPO group, although ferritin concentrations were not different among the 3 groups. Superoxide scavenging activity showed no differences. These results indicate that increased lipid peroxidation in erythrocyte, raised by decreased serum hydroxyl radical scavenging activity, is one cause of rHuEPO resistant anemia. Serum ferritin may be involved in this hydroxyl radical production.

  15. The nephroprotective properties of recombinant human erythropoietin in kidney transplantation: experimental facts and clinical proofs.

    PubMed

    Pallet, N; Rabant, M; Legendre, C; Martinez, F; Choukroun, G

    2012-12-01

    Adaptive responses to hypoxia, including hypoxia-inducible factor signaling, allow the cell to satisfy its basal metabolic demand and avoid death, but these responses can also be deleterious by promoting inflammation, cell dedifferentiation and fibrogenesis. Therefore, targeting hypoxia constitutes a promising therapeutic avenue. Recombinant human erythropoietin (rhEPO) appeared as a good candidate therapy because its hematopoietic properties could reverse anemia, and its tissue-protective properties could reduce cell death and limit maladaptive cellular responses to hypoxia. Despite experimental evidence on the nephroprotecive properties of rhEPO, recent clinical trials provided evidence that rhEPO was ineffective in preventing delayed graft function after ischemic acute injury but that the normalization of hemoglobin values preserved kidney function deterioration and reduced graft loss. Our aim here is to provide a survey of the rationale for evaluating the administration of rhEPO in the setting of kidney transplantation. We will discuss the intriguing findings that emerged from the clinical trials and the discrepancies between promising experimental results and negative clinical studies, as well as the differences in terms of the benefits and safety profiles of the normalization of hemoglobin values in chronic kidney disease patients and kidney transplant patients. PMID:23057777

  16. Delivery of erythropoietin with a needleless injection system during hemodialysis maintains plasma levels.

    PubMed

    Petersen, J; Jani, A

    2001-01-01

    Recent Occupational Safety and Health Administration guidelines require the implementation of needleless systems in dialysis units in the state of California. It is not known whether needleless systems deliver medication as well as traditional low dead-space needle/syringes. The purpose of this study was to compare the delivery of recombinant human erythropoietin (rHuEpo) by a needleless system (Medic Plastic Anti-Stick Needle/Connector, Medisystems, Seattle, WA) with delivery using a traditional low dead-space needle/syringe. We also studied the Medisystems High Flow venous chamber with integral injection site that is designed to prevent the formation of a separate serum layer within which rHuEpo may become trapped. There was no significant difference in the mean serum rHuEpo concentration achieved by either the needleless system, or traditional low dead-space needle/syringe, whether using the venous chamber's integral injection site or the postvenous chamber in-line injection site. This study demonstrates for the first time that a needleless system of delivery can reliably achieve serum rHuEpo comparable to traditional low dead-space needle/syringes.

  17. Erythropoietin promotes breast tumorigenesis through tumor-initiating cell self-renewal.

    PubMed

    Zhou, Bing; Damrauer, Jeffrey S; Bailey, Sean T; Hadzic, Tanja; Jeong, Youngtae; Clark, Kelly; Fan, Cheng; Murphy, Laura; Lee, Cleo Y; Troester, Melissa A; Miller, C Ryan; Jin, Jian; Darr, David; Perou, Charles M; Levine, Ross L; Diehn, Maximilian; Kim, William Y

    2014-02-01

    Erythropoietin (EPO) is a hormone that induces red blood cell production. In its recombinant form, EPO is the one of most prescribed drugs to treat anemia, including that arising in cancer patients. In randomized trials, EPO administration to cancer patients has been associated with decreased survival. Here, we investigated the impact of EPO modulation on tumorigenesis. Using genetically engineered mouse models of breast cancer, we found that EPO promoted tumorigenesis by activating JAK/STAT signaling in breast tumor-initiating cells (TICs) and promoted TIC self renewal. We determined that EPO was induced by hypoxia in breast cancer cell lines, but not in human mammary epithelial cells. Additionally, we demonstrated that high levels of endogenous EPO gene expression correlated with shortened relapse-free survival and that pharmacologic JAK2 inhibition was synergistic with chemotherapy for tumor growth inhibition in vivo. These data define an active role for endogenous EPO in breast cancer progression and breast TIC self-renewal and reveal a potential application of EPO pathway inhibition in breast cancer therapy. PMID:24435044

  18. HPLC-MS/MS investigation of biochemical markers for the disclosure of erythropoietin abuse in sports

    NASA Astrophysics Data System (ADS)

    Appolonova, S. A.; Dikunets, M. A.; Rodchenkov, G. M.

    2009-04-01

    The polypeptide hormone erythropoietin (EPO), which is a forbidden doping drug, was determined by high-performance liquid chromatography combined with tandem mass spectrometry (HPLC-MS/MS). The hypothesis about the influence of EPO on the asymmetric dimethylarginine (ADMA)-dimethylargininedime-thylaminohydrolase (DDAH)-NO-synthase system was verified. Changes in this system can serve as indirect biochemical markers of the presence of the forbidden EPO drug in the organism. In the test group, the concentrations of biochemical markers varied from 10 to 40 μg/ml for ADMA and symmetrical DMA (SDMA) and from 0.5 to 10 μg/ml for arginine and citrulline. A single intravenous administration of r-HuEPO (Epocrin, 2000 ME/day) for two volunteers reliably increased ADMA, SDMA, arginine, and citrulline concentrations to 40-270 μg/ml, 40-240μg/ml, 10-60 μg/ml, and 12-140 μg/ml, respectively, with respect to the reference values. The simultaneous increase in arginine, methylarginines, and citrulline contents could be an indirect marker of EPO abuse. The method is recommended for fast screening analysis.

  19. Molecular and functional characterization of erythropoietin of the goldfish (Carassius auratus L.).

    PubMed

    Katakura, Fumihiko; Katzenback, Barbara A; Belosevic, Miodrag

    2013-06-01

    Erythropoietin is the principal regulator of erythropoiesis and promotes the survival, proliferation and differentiation of erythroid progenitor cells in mammals. In this study we report on the molecular and functional characterization of erythropoietin from the goldfish. Quantitative expression analysis of goldfish epo revealed the highest mRNA levels in heart, followed by brain, liver, spleen and kidney tissues. There was no marked change of epo expression in goldfish primary kidney macrophage cultures, as progenitor cell to macrophage development progressed, indicating that erythropoietin is not involved in monopoiesis. Recombinant goldfish erythropoietin induced proliferation of progenitor cells in a dose-dependent manner, and up-regulated the expression of erythroid transcription factors gata1 and lmo2 in progenitor cells. Furthermore, recombinant goldfish erythropoietin stimulated erythroid colony formation in a dose-dependent manner and promoted survival of erythroid progenitor cells as colony-forming cells. Our results demonstrate that the function of erythropoietin in the goldfish is similar to that of mammals and suggest a highly conserved mechanism of early erythrocyte development in lower and higher vertebrates.

  20. Is erythropoietin gene a modifier factor in amyotrophic lateral sclerosis?

    PubMed

    Ghezzi, Serena; Del Bo, Roberto; Scarlato, Marina; Nardini, Martina; Carlesi, Cecilia; Prelle, Alessandro; Corti, Stefania; Mancuso, Michelangelo; Briani, Chiara; Siciliano, Gabriele; Murri, Luigi; Bresolin, Nereo; Comi, Giacomo Pietro

    2009-05-01

    To investigate the role of erythropoietin (EPO) as genetic determinant in the susceptibility to sporadic amyotrophic lateral sclerosis (SALS). We sequenced a 259-bp region spanning the 3'hypoxia-responsive element of the EPO gene in 222 Italian SALS patients and 204 healthy subjects, matched for age and ethnic origin. No potentially causative variation was detected in SALS subjects; in addition, two polymorphic variants (namely C3434T and G3544T) showed the same genotype and haplotype frequencies in patients and controls. Conversely, a weak but significant association between G3544T and age of disease onset was observed (p=0.04). Overall, our data argue against the hypothesis of EPO as a genetic risk factor for motor neuron dysfunction, at least in Italian population. However, further studies on larger cohort of patients are needed to confirm the evidence of EPO gene as modifier factor. PMID:17888545

  1. Renal erythropoietin-producing cells in health and disease

    PubMed Central

    Souma, Tomokazu; Suzuki, Norio; Yamamoto, Masayuki

    2015-01-01

    Erythropoietin (Epo) is an indispensable erythropoietic hormone primarily produced from renal Epo-producing cells (REPs). Epo production in REPs is tightly regulated in a hypoxia-inducible manner to maintain tissue oxygen homeostasis. Insufficient Epo production by REPs causes renal anemia and anemia associated with chronic disorders. Recent studies have broadened our understanding of REPs from prototypic hypoxia-responsive cells to dynamic fibrogenic cells. In chronic kidney disease, REPs are the major source of scar-forming myofibroblasts and actively produce fibrogenic molecules, including inflammatory cytokines. Notably, myofibroblast-transformed REPs (MF-REPs) recover their original physiological properties after resolution of the disease insults, suggesting that renal anemia and fibrosis could be reversible to some extent. Therefore, understanding the plasticity of REPs will lead to the development of novel targeted therapeutics for both renal fibrosis and anemia. This review summarizes the regulatory mechanisms how hypoxia-inducible Epo gene expression is attained in health and disease conditions. PMID:26089800

  2. From neurogenesis to neuroprotection in the epilepsy: signalling by erythropoietin.

    PubMed

    Castaneda-Arellano, Rolando; Beas-Zarate, Carlos; Feria-Velasco, Alfredo I; Bitar-Alatorre, Emilio W; Rivera-Cervantes, Martha C

    2014-01-01

    Epilepsy is a disorder characterised by recurrent seizures and molecular events, including the activation of early expression genes and the post-translational modifications of functional proteins. These events lead to changes in neurogenesis, mossy fibre sprouting, network reorganisation and neuronal death. The role of these events is currently a matter of great debate, especially as they relate to protection, repair, or further brain injury. In recent years, accumulating data have supported the idea that erythropoietin (EPO) regulates biological processes including neuroprotection and neurogenesis in several diseases, such as epilepsy. This review summarises the role of EPO in some of the molecular mechanisms involved in these events that could direct a more detailed approach for its use as a therapeutic alternative in reducing epileptic seizures. PMID:24896364

  3. Erythropoietin in cancer: a dilemma in risk therapy.

    PubMed

    Cao, Yihai

    2013-04-01

    Erythropoietin (EPO) is a frequently prescribed drug for treatment of cancer-related and chemotherapy-induced anemia in cancer patients. Paradoxically, recent preclinical and clinical studies indicate that EPO could potentially accelerate tumor growth and jeopardize survival in cancer patients. In this review I critically discuss the current knowledge and broad biological functions of EPO in association with tumor growth, invasion, and angiogenesis. The emphasis is focused on discussing the complex interplay between EPO and other tumor-derived factors in angiogenesis, tumor growth, invasion, and metastasis. Understanding the multifarious functions of EPO and its reciprocal relation with other signaling pathways is crucial for developing more effective agents for cancer therapy and for minimizing risks for cancer patients.

  4. Erythropoietin action in stress response, tissue maintenance and metabolism.

    PubMed

    Zhang, Yuanyuan; Wang, Li; Dey, Soumyadeep; Alnaeeli, Mawadda; Suresh, Sukanya; Rogers, Heather; Teng, Ruifeng; Noguchi, Constance Tom

    2014-01-01

    Erythropoietin (EPO) regulation of red blood cell production and its induction at reduced oxygen tension provides for the important erythropoietic response to ischemic stress. The cloning and production of recombinant human EPO has led to its clinical use in patients with anemia for two and half decades and has facilitated studies of EPO action. Reports of animal and cell models of ischemic stress in vitro and injury suggest potential EPO benefit beyond red blood cell production including vascular endothelial response to increase nitric oxide production, which facilitates oxygen delivery to brain, heart and other non-hematopoietic tissues. This review discusses these and other reports of EPO action beyond red blood cell production, including EPO response affecting metabolism and obesity in animal models. Observations of EPO activity in cell and animal model systems, including mice with tissue specific deletion of EPO receptor (EpoR), suggest the potential for EPO response in metabolism and disease. PMID:24918289

  5. The neuronal control of hypoxic ventilation: erythropoietin and sexual dimorphism.

    PubMed

    Gassmann, Max; Tissot van Patot, Martha; Soliz, Jorge

    2009-10-01

    Using mice, we demonstrated that when oxygen supply is lowered, erythropoietin (Epo), the main regulator of red blood cell production, modulates the ventilatory response by interacting with central (brainstem) and peripheral (carotid bodies) respiratory centers. We showed that enhanced Epo levels in the brainstem increased the hypoxic ventilatory response, and that intracerebroventricular injection of an Epo antagonist (soluble Epo receptor) abolished the ventilatory acclimatization to hypoxia. More recently, we have found that the impact of Epo on ventilation occurs in a sex-dependent manner. Keeping in mind that women are less susceptible to several respiratory sicknesses and syndromes than men, we suggest that Epo plays a key role in sexually-dimorphic hypoxic ventilation. Accordingly, we foresee that Epo has a potential therapeutic use as treatment for hypoxia-associated ventilatory diseases. PMID:19845617

  6. Transplantation of erythropoietin gene-modified neural stem cells improves the repair of injured spinal cord

    PubMed Central

    Wu, Min-fei; Zhang, Shu-quan; Gu, Rui; Liu, Jia-bei; Li, Ye; Zhu, Qing-san

    2015-01-01

    The protective effects of erythropoietin on spinal cord injury have not been well described. Here, the eukaryotic expression plasmid pcDNA3.1 human erythropoietin was transfected into rat neural stem cells cultured in vitro. A rat model of spinal cord injury was established using a free falling object. In the human erythropoietin-neural stem cells group, transfected neural stem cells were injected into the rat subarachnoid cavity, while the neural stem cells group was injected with non-transfected neural stem cells. Dulbecco's modified Eagle's medium/F12 medium was injected into the rats in the spinal cord injury group as a control. At 1–4 weeks post injury, the motor function in the rat lower limbs was best in the human erythropoietin-neural stem cells group, followed by the neural stem cells group, and lastly the spinal cord injury group. At 72 hours, compared with the spinal cord injury group, the apoptotic index and Caspase-3 gene and protein expressions were apparently decreased, and the bcl-2 gene and protein expressions were noticeably increased, in the tissues surrounding the injured region in the human erythropoietin-neural stem cells group. At 4 weeks, the cavities were clearly smaller and the motor and somatosensory evoked potential latencies were remarkably shorter in the human erythropoietin-neural stem cells group and neural stem cells group than those in the spinal cord injury group. These differences were particularly obvious in the human erythropoietin-neural stem cells group. More CM-Dil-positive cells and horseradish peroxidase-positive nerve fibers and larger amplitude motor and somatosensory evoked potentials were found in the human erythropoietin-neural stem cells group and neural stem cells group than in the spinal cord injury group. Again, these differences were particularly obvious in the human erythropoietin-neural stem cells group. These data indicate that transplantation of erythropoietin gene-modified neural stem cells into the

  7. Transplantation of erythropoietin gene-modified neural stem cells improves the repair of injured spinal cord.

    PubMed

    Wu, Min-Fei; Zhang, Shu-Quan; Gu, Rui; Liu, Jia-Bei; Li, Ye; Zhu, Qing-San

    2015-09-01

    The protective effects of erythropoietin on spinal cord injury have not been well described. Here, the eukaryotic expression plasmid pcDNA3.1 human erythropoietin was transfected into rat neural stem cells cultured in vitro. A rat model of spinal cord injury was established using a free falling object. In the human erythropoietin-neural stem cells group, transfected neural stem cells were injected into the rat subarachnoid cavity, while the neural stem cells group was injected with non-transfected neural stem cells. Dulbecco's modified Eagle's medium/F12 medium was injected into the rats in the spinal cord injury group as a control. At 1-4 weeks post injury, the motor function in the rat lower limbs was best in the human erythropoietin-neural stem cells group, followed by the neural stem cells group, and lastly the spinal cord injury group. At 72 hours, compared with the spinal cord injury group, the apoptotic index and Caspase-3 gene and protein expressions were apparently decreased, and the bcl-2 gene and protein expressions were noticeably increased, in the tissues surrounding the injured region in the human erythropoietin-neural stem cells group. At 4 weeks, the cavities were clearly smaller and the motor and somatosensory evoked potential latencies were remarkably shorter in the human erythropoietin-neural stem cells group and neural stem cells group than those in the spinal cord injury group. These differences were particularly obvious in the human erythropoietin-neural stem cells group. More CM-Dil-positive cells and horseradish peroxidase-positive nerve fibers and larger amplitude motor and somatosensory evoked potentials were found in the human erythropoietin-neural stem cells group and neural stem cells group than in the spinal cord injury group. Again, these differences were particularly obvious in the human erythropoietin-neural stem cells group. These data indicate that transplantation of erythropoietin gene-modified neural stem cells into the

  8. Nonclinical evaluation of the potential for mast cell activation by an erythropoietin analog

    SciTech Connect

    Weaver, James L.

    2015-09-15

    The erythropoietin analog peginesatide was withdrawn from marketing due to unexpected severe anaphylactic reactions associated with administration of the multi-use formulation. The adverse events occurred rapidly following the first ever administration of the drug with most affected patients becoming symptomatic in less than 30 min. This is most consistent with an anaphylactoid reaction due to direct activation of mast cells. Laboratory evaluation was undertaken using rat peritoneal mast cells as the model system. Initial studies showed that high concentrations of the formulated drug as well as formulated vehicle alone could cause mast cell degranulation as measured by histamine release. The purified active drug was not able to cause histamine release whereas the vehicle filtrate and lab created drug vehicle were equally potent at causing histamine release. Individual formulations of vehicle leaving one component out showed that histamine release was due to phenol. Dose response studies with phenol showed a very sharp dose response curve that was similar in three buffer systems. Cellular analysis by flow cytometry showed that the histamine release was not due to cell death, and that changes in light scatter parameters consistent with degranulation were rapidly observed. Limited testing with primary human mast cells showed a similar dose response of histamine release with exposure to phenol. To provide in vivo confirmation, rats were injected with vehicle formulated with various concentrations of phenol via a jugular vein cannula. Significant release of histamine was detected in blood samples taken 2 min after dosing at the highest concentrations tested. - Highlights: • Peginesatide caused severe anaphylactoid reactions in 0.2% of patients. • Both formulated drug and vehicle cause degranulation of rat mast cells. • Phenol was identified as the vehicle component causing degranulation. • Human mast cells show similar dose response to phenol as rat mast cells

  9. Erythropoietin critically regulates the terminal maturation of murine and human primitive erythroblasts

    PubMed Central

    Malik, Jeffrey; Kim, Ah Ram; Tyre, Kaitlin A.; Cherukuri, Anjuli R.; Palis, James

    2013-01-01

    Primitive erythroid cells, the first red blood cells produced in the mammalian embryo, are necessary for embryonic survival. Erythropoietin and its receptor EpoR, are absolutely required for survival of late-stage definitive erythroid progenitors in the fetal liver and adult bone marrow. Epo- and Epor-null mice die at E13.5 with a lack of definitive erythrocytes. However, the persistence of circulating primitive erythroblasts raises questions about the role of erythropoietin/EpoR in primitive erythropoiesis. Using Epor-null mice and a novel primitive erythroid 2-step culture we found that erythropoietin is not necessary for specification of primitive erythroid progenitors. However, Epor-null embryos develop a progressive, profound anemia by E12.5 as primitive erythroblasts mature as a synchronous cohort. This anemia results from reduced primitive erythroblast proliferation associated with increased p27 expression, from advanced cellular maturation, and from markedly elevated rates of apoptosis associated with an imbalance in pro- and anti-apoptotic gene expression. Both mouse and human primitive erythroblasts cultured without erythropoietin also undergo accelerated maturation and apoptosis at later stages of maturation. We conclude that erythropoietin plays an evolutionarily conserved role in promoting the proliferation, survival, and appropriate timing of terminal maturation of primitive erythroid precursors. PMID:23894012

  10. Bone marrow mesenchymal stem cells transplantation promotes the release of endogenous erythropoietin after ischemic stroke

    PubMed Central

    Lv, Wen; Li, Wen-yu; Xu, Xiao-yan; Jiang, Hong; Bang, Oh Yong

    2015-01-01

    This study investigated whether bone marrow mesenchymal stem cell (BMSC) transplantation protected ischemic cerebral injury by stimulating endogenous erythropoietin. The model of ischemic stroke was established in rats through transient middle cerebral artery occlusion. Twenty-four hours later, 1 × 106 human BMSCs (hBMSCs) were injected into the tail vein. Fourteen days later, we found that hBMSCs promoted the release of endogenous erythropoietin in the ischemic region of rats. Simultaneously, 3 μg/d soluble erythropoietin receptor (sEPOR) was injected into the lateral ventricle, and on the next 13 consecutive days. sEPOR blocked the release of endogenous erythropoietin. The neurogenesis in the subventricular zone was less in the hBMSCs + sEPOR group than in the hBMSCs + heat-denatured sEPOR group. The adhesive-removal test result and the modified Neurological Severity Scores (mNSS) were lower in the hBMSCs + sEPOR group than in the heat-denatured sEPOR group. The adhesive-removal test result and mNSS were similar between the hBMSCs + heat-denatured sEPOR group and the hBMSCs + sEPOR group. These findings confirm that BMSCs contribute to neurogenesis and improve neurological function by promoting the release of endogenous erythropoietin following ischemic stroke. PMID:26487854

  11. The hematopoietic growth factor "erythropoietin" enhances the therapeutic effect of mesenchymal stem cells in Alzheimer's disease.

    PubMed

    Khairallah, M I; Kassem, L A; Yassin, N A; El Din, M A Gamal; Zekri, M; Attia, M

    2014-01-01

    Alzheimer's disease is a neurodegenerative disorder clinically characterized by cognitive dysfunction and by deposition of amyloid plaques, neurofibrillary tangles in the brain. The study investigated the therapeutic effect of combined mesenchymal stem cells and erythropoietin on Alzheimer's disease. Five groups of mice were used: control group, Alzheimer's disease was induced in four groups by a single intraperitoneal injection of 0.8 mg kg(-1) lipopolysaccharide and divided as follows: Alzheimer's disease group, mesenchymal stem cells treated group by injecting mesenchymal stem cells into the tail vein (2 x 10(6) cells), erythropoietin treated group (40 microg kg(-1) b.wt.) injected intraperitoneally 3 times/week for 5 weeks and mesenchymal stem cells and erythropoietin treated group. Locomotor activity and memory were tested using open field and Y-maze. Histological, histochemical, immunohistochemical studies, morphometric measurements were examined in brain sections of all groups. Choline transferase activity, brain derived neurotrophic factor expression and mitochondrial swellings were assessed in cerebral specimens. Lipopolysaccharide decreased locomotor activity, memory, choline transferase activity and brain derived neurotrophic factor. It increased mitochondrial swelling, apoptotic index and amyloid deposition. Combined mesenchymal stem cells and erythropoietin markedly improved all these parameters. This study proved the effective role of mesenchymal stem cells in relieving Alzheimer's disease symptoms and manifestations; it highlighted the important role of erythropoietin in the treatment of Alzheimer's disease.

  12. Alternative erythropoietin-mediated signaling prevents secondary microvascular thrombosis and inflammation within cutaneous burns.

    PubMed

    Bohr, Stefan; Patel, Suraj J; Shen, Keyue; Vitalo, Antonia G; Brines, Michael; Cerami, Anthony; Berthiaume, Francois; Yarmush, Martin L

    2013-02-26

    Alternate erythropoietin (EPO)-mediated signaling via the heteromeric receptor composed of the EPO receptor and the β-common receptor (CD131) exerts the tissue-protective actions of EPO in various types of injuries. Herein we investigated the effects of the EPO derivative helix beta surface peptide (synonym: ARA290), which specifically triggers alternate EPO-mediated signaling, but does not bind the erythropoietic EPO receptor homodimer, on the progression of secondary tissue damage following cutaneous burns. For this purpose, a deep partial thickness cutaneous burn injury was applied on the back of mice, followed by systemic administration of vehicle or ARA290 at 1, 12, and 24 h postburn. With vehicle-only treatment, wounds exhibited secondary microvascular thrombosis within 24 h postburn, and subsequent necrosis of the surrounding tissue, thus converting to a full-thickness injury within 48 h. On the other hand, when ARA290 was systemically administered, patency of the microvasculature was maintained. Furthermore, ARA290 mitigated the innate inflammatory response, most notably tumor necrosis factor-alpha-mediated signaling. These findings correlated with long-term recovery of initially injured yet viable tissue components. In conclusion, ARA290 may be a promising therapeutic approach to prevent the conversion of partial- to full-thickness burn injuries. In a clinical setting, the decrease in burn depth and area would likely reduce the necessity for extensive surgical debridement as well as secondary wound closure by means of skin grafting. This use of ARA290 is consistent with its tissue-protective properties previously reported in other models of injury, such as myocardial infarction and hemorrhagic shock. PMID:23401545

  13. EPC mobilization after erythropoietin treatment in acute ST-elevation myocardial infarction: the REVEAL EPC substudy

    PubMed Central

    Povsic, Thomas J.; Najjar, Samer S.; Prather, Kristi; Zhou, Jiying; Adams, Stacie D.; Zavodni, Katherine L.; Kelly, Francine; Melton, Laura G.; Hasselblad, Vic; Heitner, John F.; Raman, Subha V.; Barsness, Gregory W.; Patel, Manesh R.; Kim, Raymond J.; Lakatta, Edward G.; Harrington, Robert A.; Rao, Sunil V.

    2014-01-01

    Erythropoietin (EPO) was hypothesized to mitigate reperfusion injury, in part via mobilization of endothelial progenitor cells (EPCs). The REVEAL trial found no reduction in infarct size with a single dose of EPO (60,000 U) in patients with ST-segment elevation myocardial infarction. In a substudy, we aimed to determine the feasibility of cryopreserving and centrally analyzing EPC levels to assess the relationship between EPC numbers, EPO administration, and infarct size. As a prespecified substudy, mononuclear cells were locally cryopreserved before as well as 24 and 48–72 h after primary percutaneous coronary intervention. EPC samples were collected in 163 of 222 enrolled patients. At least one sample was obtained from 125 patients, and all three time points were available in 83 patients. There were no significant differences in the absolute EPC numbers over time or between EPO- and placebo-treated patients; however, there was a trend toward a greater increase in EPC levels from 24 to 48–72 h postintervention in patients receiving ≥30,000 U of EPO (P = 0.099 for CD133+ cells, 0.049 for CD34+ cells, 0.099 for ALDHbr cells). EPC numbers at baseline were inversely related to infarct size (P = 0.03 for CD133+ cells, 0.006 for CD34+ cells). Local whole cell cryopreservation and central EPC analysis in the context of a multicenter randomized trial is feasible but challenging. High-dose (≥30,000 U) EPO may mobilize EPCs at 48–72 h, and baseline EPC levels may be inversely associated with infarct size. PMID:23700090

  14. Protective effects of BMSCs in combination with erythropoietin in bronchopulmonary dysplasia-induced lung injury.

    PubMed

    Zhang, Zhao-Hua; Pan, Yan-Yan; Jing, Rui-Sheng; Luan, Yun; Zhang, Luan; Sun, Chao; Kong, Feng; Li, Kai-Lin; Wang, Yi-Biao

    2016-08-01

    Bronchopulmonary dysplasia (BPD) is the most common type of chronic lung disease in infancy, for which no effective therapy is currently available. The aim of the present study was to investigate the effect of treatment with bone marrow mesenchymal stem cells (BMSCs) in combination with recombinant human erythropoietin (rHuEPO) on BPD‑induced mouse lung injury, and discuss the underlying mechanism. The BPD model was established by the exposure of neonatal mice to continuous high oxygen exposure for 14 days, following which 1x106 BMSCs and 5,000 U/kg rHuEPO were injected into the mice 1 h prior to and 7 days following exposure to hyperoxia. The animals received four treatments in total (n=10 in each group). After 14 days, the body weights, airway structure, and levels of matrix metalloproteinase‑9 (MMP‑9) and vascular endothelial growth factor (VEGF) were detected using histological and immunohistochemical analyses. The effect on cell differentiation was observed by examining the presence of platelet endothelial cell adhesion molecule (PECAM) and VEGF using immunofluorescence. Compared with the administration of BMSCs alone, the body weight, airway structure, and the levels of MMP‑9 and VEGF were significantly improved in the BMSCs/rHuEPO group. The results of the present study demonstrated that the intravenous injection of BMSCs significantly improved lung damage in the hyperoxia‑exposed neonatal mouse model. Furthermore, the injection of BMSCs in combination with intraperitoneal injection of rHuEPO had a more marked effect, compared with BMSCs alone, and the mechanism may be mediated by the promoting effects of BMSCs and EPO. The results of the present study provided information, which may assist in future clinical trials. PMID:27279073

  15. Evaluation of bioactivity and pharmacokinetic characteristics of PEGylated P.pastoris-expressed erythropoietin.

    PubMed

    Maleki, Ahmad; Najafabadi, Abdolhossein Rouholamini; Roohvand, Farzin; Shafiee, Abbas; Khanahmad, Hossein; Faghihi, Homa; Hedayati, Mohammad H; Tajerzadeh, Hosnieh

    2011-11-01

    High costs of production and relatively short serum half-life of mammalian cell-derived recombinant human erythropoietin (rHuEpo) necessitate finding and developing superior hosts/technologies for more efficient production of longer-acting erythropoietic agents. With these aims, we provide the first report on reductive alkylation of low-cost P.pastoris-expressed rHuEpo (PPEpo) with PEG-aldehyde. The PCR-amplified cDNA of native rHuEpo was cloned into the pPICZαA vector and transformed into the yeast Pichia pastoris. The best expressing transformant was selected and employed for secreted-expression of PPEpo using the standard protocols. Purified PPEpo was N-terminally PEGylated with 20-kDa mPEG-propionaldehyde in a low pH (5) condition. The in vitro and in vivo biological activities of purified mono-PEGylated PPEpo was evaluated by the UT-7 cells proliferation assay and normocythaemic mice assay, respectively. Pharmacokinetic parameters were determined following intravenous administration of Epo proteins in rabbits. While PPEpo showed a higher in vitro bioactivity compared to rHuEpo, no in vivo efficiency was determined for PPEpo. However, the in vivo activity of PEG-PPEpo conjugate was comparable to that of rHuEpo. Pharmacokinetic studies showed that the terminal half-life and mean residence time of PEG-PPEpo were increased approximately 4-fold and 6.5-fold respectively, compared with those of PPEpo. The results indicate that N-terminal PEGylation of Pichia-expressed Epo could be considered as a promising approach for generating cost-effective and long-acting erythropoiesis-stimulating agents.

  16. Erythropoietin research, 80 years after the initial studies by Carnot and Deflandre.

    PubMed

    Jelkmann, W

    1986-03-01

    The discovery and present state of knowledge of the specific hormonal mechanism controlling erythropoiesis are briefly reviewed. 80 years ago, Carnot and Deflandre postulated the existence of a humoral erythropoietic factor ('hémopoïétine') produced in response to anemia. The general approval of this concept required almost 50 years, when the factor was recognized to be a glycoprotein hormone and termed 'erythropoietin'. Very recently, human erythropoietin has been purified and its amino acid sequence been identified. The hormone is mainly of renal origin. Its production is stimulated by tissue hypoxia of different causes. The mechanism of O2 sensing and the exact site of erythropoietin synthesis in the kidney still remain to be clarified.

  17. Towards erythropoietin equations that estimate oxygen delivery rather than static hemoglobin targets.

    PubMed

    Diskin, Charles J

    2012-01-01

    Although we have known since the 19th century that oxygen tension affects erythrocyte production, we have only recently begun to understand many subtleties of erythropoietin physiology. The unanticipated increase in mortality associated with erythropoietin use found in recent randomized studies is prompting a reassessment of static hemoglobin targets. Hemoglobin levels in dialysis patients do not correlate with endogenous erythropoietin production and may be related to differences in oxygen delivery resulting from shifts in the oxygen-hemoglobin dissociation curve. The time may have arrived to develop more physiologic targets such as oxygen delivery that would mimic the natural response to hypoxia. There are several equations that already exist that can compensate for the effects of the concentration of inorganic and organic phosphates as well as pH, carbon dioxide, and temperature on the delivery of oxygen. However, since the shape and dispersion of the oxygen-hemoglobin dissociation curve may actually change in different disease states, more work is needed.

  18. The anemia of microgravity and recumbency. Role of sympathetic neural control of erythropoietin production

    NASA Astrophysics Data System (ADS)

    Robertson, David; Krantz, Sanford B.; Biaggioni, Italo

    We hypothesize that reduced sympathetic stimulation of erythropoietin production may maintain the anemia which develops in virtually all space travellers. We tested this hypothesis in a human model of reduced sympathetic activity. Thirty-three patients with the Bradbury-Eggleston syndrome were divided into three groups according to their hemoglobin (Hgb) level. Patients with low Hgb had lower upright norepinephrine and lower upright renin. Patients with anemia also had inappropriately low plasma erythropoietin levels. We administered recombinant erythropoietin (Epogen) 25-50 units/kg s.c. 3 times per week and found that the anemia seen in autonomic failure could be reversed by this treatment. These results support the hypothesis that erythropoiesis is modulated by the sympathetic nervous system and that such mechanisms may also operate in the microgravity environment where sympathetic activity is reduced.

  19. Erythropoietin and interleukin-2 activate distinct JAK kinase family members.

    PubMed Central

    Barber, D L; D'Andrea, A D

    1994-01-01

    The erythropoietin (EPO) receptor and the interleukin-2 (IL-2) receptor beta-chain subunit are members of the cytokine receptor superfamily. They have conserved primary amino acid sequences in their cytoplasmic domains and activate phosphorylation of common substrates, suggesting common biochemical signaling mechanisms. We have generated a cell line, CTLL-EPO-R, that contains functional cell surface receptors for both EPO and IL-2. CTLL-EPO-R cells demonstrated similar growth kinetics in EPO and IL-2. Stimulation with EPO resulted in the rapid, dose-dependent tyrosine phosphorylation of JAK2. In contrast, stimulation with IL-2 or the related cytokine IL-4 resulted in the rapid, dose-dependent tyrosine phosphorylation of JAK1 and an additional 116-kDa protein. This 116-kDa protein was itself immunoreactive with a polyclonal antiserum raised against JAK2 and appears to be a novel member of the JAK kinase family. Immune complex kinase assays confirmed that IL-2 and IL-4 activated JAK1 and EPO activated JAK2. These results demonstrate that multiple biochemical pathways are capable of conferring a mitogenic signal in CTLL-EPO-R cells and that the EPO and IL-2 receptors interact with distinct JAK kinase family members within the same cellular background. Images PMID:7935373

  20. Murine erythropoietin gene: cloning, expression, and human gene homology.

    PubMed Central

    Shoemaker, C B; Mitsock, L D

    1986-01-01

    The gene for murine erythropoietin (EPO) was isolated from a mouse genomic library with a human EPO cDNA probe. Nucleotide sequence analysis permitted the identification of the murine EPO coding sequence and the prediction of the encoded amino acid sequence based on sequence conservation between the mouse and human EPO genes. Both the coding DNA and the amino acid sequences were 80% conserved between the two species. Transformation of COS-1 cells with a mammalian cell expression vector containing the murine EPO coding region resulted in secretion of murine EPO with biological activity on both murine and human erythroid progenitor cells. The transcription start site for the murine EPO gene in kidneys was determined. This permitted tentative identification of the transcription control region. The region included 140 base pairs upstream of the cap site which was over 90% conserved between the murine and human genes. Surprisingly, the first intron and much of the 5'- and 3'-untranslated sequences were also substantially conserved between the genes of the two species. Images PMID:3773894

  1. Erythropoietin regulates Treg cells in asthma through TGFβ receptor signaling.

    PubMed

    Wan, Guoshi; Wei, Bing

    2015-01-01

    Asthma is a chronic inflammatory disorder of the airways, the development of which is suppressed by regulatory T cells (Treg). Erythropoietin (EPO) is originally defined as a hematopoietic growth factor. Recently, the anti-inflammatory effects of EPO in asthma have been acknowledged. However, the underlying mechanisms remain ill-defined. Here, we showed that EPO treatment significantly reduced the severity of an ovalbumin (OVA)-induced asthma in mice, seemingly through promoting Foxp3-mediated activation of Treg cells in OVA-treated mouse lung. The activation of Treg cells resulted from increases in transforming growth factor β1 (TGFβ1), which were mainly produced by M2 macrophages (M2M). In vitro, Co-culture with M2M increased Foxp3 levels in Treg cells and the Treg cell number, in a TGFβ receptor signaling dependent manner. Moreover, elimination of macrophages abolished the therapeutic effects of EPO in vivo. Together, our data suggest that EPO may increase M2M, which activate Treg cells through TGFβ receptor signaling to mitigate the severity of asthma.

  2. Erythropoietin regulates Treg cells in asthma through TGFβ receptor signaling

    PubMed Central

    Wan, Guoshi; Wei, Bing

    2015-01-01

    Asthma is a chronic inflammatory disorder of the airways, the development of which is suppressed by regulatory T cells (Treg). Erythropoietin (EPO) is originally defined as a hematopoietic growth factor. Recently, the anti-inflammatory effects of EPO in asthma have been acknowledged. However, the underlying mechanisms remain ill-defined. Here, we showed that EPO treatment significantly reduced the severity of an ovalbumin (OVA)-induced asthma in mice, seemingly through promoting Foxp3-mediated activation of Treg cells in OVA-treated mouse lung. The activation of Treg cells resulted from increases in transforming growth factor β1 (TGFβ1), which were mainly produced by M2 macrophages (M2M). In vitro, Co-culture with M2M increased Foxp3 levels in Treg cells and the Treg cell number, in a TGFβ receptor signaling dependent manner. Moreover, elimination of macrophages abolished the therapeutic effects of EPO in vivo. Together, our data suggest that EPO may increase M2M, which activate Treg cells through TGFβ receptor signaling to mitigate the severity of asthma. PMID:26807178

  3. Physiological focus on the erythropoietin-hepcidin-ferroportin axis.

    PubMed

    D'Anna, María Cecilia; Roque, Marta Elena

    2013-05-01

    To analyze the interconnection between erythropoiesis and iron metabolism, one of the issues raised in this study was to know iron bioavailability under physiopathological conditions. Our aim was to understand the functional axis response composed of erythropoietin (Epo)-hepcidin-ferroportin (FPN), when 2 dysfunctional states coexist, using an animal model of iron overload followed by hypoxia. FPN and prohepcidin were assessed by immunohistochemistry using rabbit anti-mouse FPN polyclonal and prohepcidin monoclonal antibodies. Goat-labeled polymer - horseradish peroxidase anti-rabbit EnVision + System (DAB) was used as the secondary antibody. Epo levels were measured by ELISA. Tissue iron was studied by Prussian blue iron staining. Erythropoietic response was assessed using conventional hematological tests. Iron overload increased prohepcidin that remained high in hypoxia, coexisting with high levels of Epo in hypoxia, with or without iron overload. In hypoxia, FPN was clearly evident in reticuloendothelial macrophages, more than in hypoxia with iron overload. Interestingly, duodenal FPN was clearly identified on the basolateral membrane in hypoxia, with or without iron overload. Our data indicate that 2 signals could induce the cell-specific response as follows: (i) iron signal, induced prohepcidin, which reduced reticuloendothelial FPN and reduced iron availability; and (ii) hypoxia signal, stimulated Epo, which affected iron absorption by stabilizing duodenal FPN and allowed iron supply to erythropoiesis independently of store size.

  4. Erythropoietin: a novel therapy for hypoxic-ischaemic encephalopathy?

    PubMed

    Wu, Yvonne W; Gonzalez, Fernando F

    2015-04-01

    Perinatal hypoxic-ischaemic encephalopathy (HIE) occurs in 1 to 3 per 1000 term births. HIE is not preventable in most cases, and therapies are limited. Hypothermia improves outcomes and is the current standard of care. Yet, clinical trials suggest that 44-53% of infants who receive hypothermia will die or suffer moderate to severe neurological disability. In this article, we review the preclinical and clinical evidence for erythropoietin (EPO) as a potential novel neuroprotective agent for the treatment of HIE. EPO is a novel neuroprotective agent, with remarkable neuroprotective and neuroregenerative effects in animals. Rodent and primate models of neonatal brain injury support the safety and efficacy of multiple EPO doses for improving histological and functional outcomes after hypoxia-ischaemia. Small clinical trials of EPO in neonates with HIE have also provided evidence supporting safety and preliminary efficacy in humans. There is currently insufficient evidence to support the use of high-dose EPO in newborns with HIE. However, several on-going trials will provide much needed data regarding the safety and efficacy of this potential new therapy when given in conjunction with hypothermia for HIE. Novel neuroprotective therapies are needed to further reduce the rate and severity of neurodevelopmental disabilities resulting from HIE. High-dose EPO is a promising therapy that can be administered in conjunction with hypothermia. However, additional data are needed to determine the safety and efficacy of this adjuvant therapy for HIE.

  5. Erythropoietin: a novel therapy for hypoxic-ischaemic encephalopathy?

    PubMed

    Wu, Yvonne W; Gonzalez, Fernando F

    2015-04-01

    Perinatal hypoxic-ischaemic encephalopathy (HIE) occurs in 1 to 3 per 1000 term births. HIE is not preventable in most cases, and therapies are limited. Hypothermia improves outcomes and is the current standard of care. Yet, clinical trials suggest that 44-53% of infants who receive hypothermia will die or suffer moderate to severe neurological disability. In this article, we review the preclinical and clinical evidence for erythropoietin (EPO) as a potential novel neuroprotective agent for the treatment of HIE. EPO is a novel neuroprotective agent, with remarkable neuroprotective and neuroregenerative effects in animals. Rodent and primate models of neonatal brain injury support the safety and efficacy of multiple EPO doses for improving histological and functional outcomes after hypoxia-ischaemia. Small clinical trials of EPO in neonates with HIE have also provided evidence supporting safety and preliminary efficacy in humans. There is currently insufficient evidence to support the use of high-dose EPO in newborns with HIE. However, several on-going trials will provide much needed data regarding the safety and efficacy of this potential new therapy when given in conjunction with hypothermia for HIE. Novel neuroprotective therapies are needed to further reduce the rate and severity of neurodevelopmental disabilities resulting from HIE. High-dose EPO is a promising therapy that can be administered in conjunction with hypothermia. However, additional data are needed to determine the safety and efficacy of this adjuvant therapy for HIE. PMID:25800490

  6. Study on the inflammatory intervention of erythropoietin on NEC

    PubMed Central

    QI, WEIBIN; SHEN, QIONG; ZHANG, LIN; HAN, LI-PING; WANG, SUMIN

    2016-01-01

    The aim of this study was to investigate the effect of erythropoietin (EPO) on the inflammatory response and the mechanism analysis of the Τoll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling pathway of NEC. A total of 94 patients with necrotizing enterocolitis (NEC) were randomly divided into the control (42 cases) and observation (52 cases) groups, The control group received the standard medical treatment plan, whereas for the observation group this treatment plan was combined with the application of recombinant EPO for intramuscular injection treatment. The clinical effect was subsequently compared. The results showed that the complication and death rates in the observation group were significantly lower than those in the control group with statistically significant differences (P<0.05). Following treatments, the levels of tumor necrosis factor-α (TNF-α) and interleukin (IL)-6 in the observation group were significantly lower than those in the control group. The expression levels of mRNA of TLR4 and NF-κB in the observation group were significantly lower than those in the control group, with statistically significant differences (P<0.05). In summary, EPO was able to reduce the levels of inflammatory response of TNF-α and IL-6 through the TLR4/NF-κB signaling pathway, and improve the NEC, thus providing a basis for the clinical treatment of NEC. PMID:27284304

  7. Double blind trial of recombinant human erythropoietin in preterm infants.

    PubMed Central

    Emmerson, A J; Coles, H J; Stern, C M; Pearson, T C

    1993-01-01

    Twenty four infants between 27 and 33 weeks' gestation were recruited into a double blind study to investigate the use of recombinant human erythropoietin (r-HuEpo) for the prevention of anaemia of prematurity. Between 50 and 150 U of r-HuEpo (n = 16) or placebo was administered subcutaneously twice a week from 7 days of age until discharge. There was a significant increase in the reticulocyte count in infants receiving r-HuEpo sustained from the second week of treatment until discharge compared with placebo. There was a reduction in the number of transfusions required in the r-HuEpo group with only 47% requiring a transfusion compared with 87% in the placebo group. During treatment with r-HuEpo there was a significant rise in the red cell folate concentration, a significant fall in the ferritin concentration, and a significantly higher percentage of haemoglobin F at discharge suggesting active erythropoiesis. The study provides strong evidence for the efficacy of r-HuEpo in stimulating erythropoiesis and reducing the requirement for transfusions for anaemia of prematurity. PMID:8466265

  8. Hypoxia Signaling Cascade for Erythropoietin Production in Hepatocytes.

    PubMed

    Tojo, Yutaka; Sekine, Hiroki; Hirano, Ikuo; Pan, Xiaoqing; Souma, Tomokazu; Tsujita, Tadayuki; Kawaguchi, Shin-ichi; Takeda, Norihiko; Takeda, Kotaro; Fong, Guo-Hua; Dan, Takashi; Ichinose, Masakazu; Miyata, Toshio; Yamamoto, Masayuki; Suzuki, Norio

    2015-08-01

    Erythropoietin (Epo) is produced in the kidney and liver in a hypoxia-inducible manner via the activation of hypoxia-inducible transcription factors (HIFs) to maintain oxygen homeostasis. Accelerating Epo production in hepatocytes is one plausible therapeutic strategy for treating anemia caused by kidney diseases. To elucidate the regulatory mechanisms of hepatic Epo production, we analyzed mouse lines harboring liver-specific deletions of genes encoding HIF-prolyl-hydroxylase isoforms (PHD1, PHD2, and PHD3) that mediate the inactivation of HIF1α and HIF2α under normal oxygen conditions. The loss of all PHD isoforms results in both polycythemia, which is caused by Epo overproduction, and fatty livers. We found that deleting any combination of two PHD isoforms induces polycythemia without steatosis complications, whereas the deletion of a single isoform induces no apparent phenotype. Polycythemia is prevented by the loss of either HIF2α or the hepatocyte-specific Epo gene enhancer (EpoHE). Chromatin analyses show that the histones around EpoHE dissociate from the nucleosome structure after HIF2α activation. HIF2α also induces the expression of HIF3α, which is involved in the attenuation of Epo production. These results demonstrate that the total amount of PHD activity is more important than the specific function of each isoform for hepatic Epo expression regulated by a PHD-HIF2α-EpoHE cascade in vivo.

  9. Erythropoietin and Cancer: The Unintended Consequences of Anemia Correction

    PubMed Central

    Debeljak, Nataša; Solár, Peter; Sytkowski, Arthur J.

    2014-01-01

    Until 1990, erythropoietin (EPO) was considered to have a single biological purpose and action, the stimulation of red blood cell growth and differentiation. Slowly, scientific and medical opinion evolved, beginning with the discovery of an effect on endothelial cell growth in vitro and the identification of EPO receptors (EPORs) on neuronal cells. We now know that EPO is a pleiotropic growth factor that exhibits an anti-apoptotic action on numerous cells and tissues, including malignant ones. In this article, we present a short discussion of EPO, receptors involved in EPO signal transduction, and their action on non-hematopoietic cells. This is followed by a more detailed presentation of both pre-clinical and clinical data that demonstrate EPO’s action on cancer cells, as well as tumor angiogenesis and lymphangiogenesis. Clinical trials with reported adverse effects of chronic erythropoiesis-stimulating agents (ESAs) treatment as well as clinical studies exploring the prognostic significance of EPO and EPOR expression in cancer patients are reviewed. Finally, we address the use of EPO and other ESAs in cancer patients. PMID:25426117

  10. Antioxidants may Attenuate Plasma Erythropoietin Decline after Hyperbaric Oxygen Diving.

    PubMed

    Mutzbauer, T S; Schneider, M; Neubauer, B; Weiss, M; Tetzlaff, K

    2015-11-01

    According to previous studies, plasma erythropoietin (EPO) may decrease after hyperbaric oxygen exposure due to oxidative stress. It is hypothesized that the decrease of EPO can be attenuated by oxygen free radical scavengers.The aim of the present study was to evaluate whether EPO plasma levels can be influenced by oral application of vitamin C and E before repeated hyperbaric oxygen exposure during diving. 16 healthy male police task force divers performed 3 morning dives on oxygen within a regular diving schedule on 3 consecutive days. They were randomized into either the placebo group or the vitamin group, receiving 1 g ascorbic acid and 600 IU D-α-tocopherol orally 60 min before the dive. Blood samples for EPO measurement were taken on days 1, 2, and 3 at T1, T3 and T5 60 min before and at T2, T4 and T6 60 min after each dive, respectively. A moderate decrease of EPO was observed beginning at T3 until T6 in the placebo group. The EPO concentrations in the vitamin group did not show relevant variations compared to baseline. Radical scavenging vitamins C and D may counteract hyperbaric oxygen related mechanisms reducing EPO production in hyperbaric oxygen exposure during diving.

  11. Past, present and future of erythropoietin use in the elderly.

    PubMed

    de Francisco, Angel L M; Fernandez Fresnedo, Gema; Rodrigo, Emilio; Piñera, Celestino; Heras, Milagros; Palomar, Rosa; Ruiz, Juan C; Arias, Manuel

    2002-01-01

    More than a decade has passed since the first patient with end-stage renal failure was treated with erythropoietin (EPO) and more than 85% of patients now receive this therapy. In the year 2002 more than 60% of dialysis patients will be elderly, and the treatment of anemia will be more complex due to the aditional causes: folate, iron and vitamin deficiency in this population. Correction of anemia with EPO brings about partial regression of left ventricular hypertrophy and some data suggest that such treatment reduces cardiovascular mortality in patients without advance cardiac disease. Normalization of hematocrit with EPO increases oxygen supply to the brain tissue with improvement in brain function. The improvement in the ability to recognize, discriminate and hold stimuli in memory for difficult tasks is particularly important for elderly people. No differences have been noted in the incidence of clotting of vascular access in patients treated with EPO compared with hemodialysis patients not so treated. Also no one has demostrated that treatment with EPO accelerates renal decline in patients with progressive renal insufficiency. In elderly people with anemia secondary to advanced renal failure, EPO therapy improves physical, cognitive and sexual function, and health related quality of life.

  12. Regulated expression of erythropoietin by two human hepatoma cell lines

    SciTech Connect

    Goldberg, M.A.; Glass, G.A.; Cunningham, J.M.; Bunn, H.F.

    1987-11-01

    The development of a cell culture system that produces erythropoietin (Epo) in a regulated manner has been the focus of much effort. The authors have screened multiple renal and hepatic cell lines for either constitutive or regulated expression of Epo. Only the human hepatoma cell lines, Hep3B and HepG2, made significant amounts of Epo as measured both by radioimmunoassay and in vitro bioassay (as much as 330 milliunits per 10/sup 6/ cells in 24 hr). The constitutive production of Epo increased dramatically as a function of cell density in both cell lines. At cell densities < 3.3 x 10/sup 5/ cells per cm/sup 2/, there was little constitutive release of Epo in the medium. With Hep3B cells grown at low cell densities, a mean 18-fold increase in Epo expression was seen in response to hypoxia and a 6-fold increase was observed in response to incubation in medium containing 50 ..mu..M cobalt(II) chloride. At similar low cell densities, Epo production in HepG2 cells could be enhanced an average of about 3-fold by stimulation with either hypoxia or cobalt(II) chloride. Upon such stimulation, both cell lines demonstrated markedly elevated levels of Epo mRNA. Hence, both Hep3B and HepG2 cell lines provide an excellent in vitro system in which to study the physiological regulation of Epo expression.

  13. Erythropoietin reduces the expression of myostatin in mdx dystrophic mice.

    PubMed

    Feder, D; Rugollini, M; Santomauro, A; Oliveira, L P; Lioi, V P; Santos, R dos; Ferreira, L G; Nunes, M T; Carvalho, M H; Delgado, P O; Carvalho, A A S; Fonseca, F L A

    2014-11-01

    Erythropoietin (EPO) has been well characterized as a renal glycoprotein hormone regulating red blood cell production by inhibiting apoptosis of erythrocyte progenitors in hematopoietic tissues. EPO exerts regulatory effects in cardiac and skeletal muscles. Duchenne muscular dystrophy is a lethal degenerative disorder of skeletal and cardiac muscle. In this study, we tested the possible therapeutic beneficial effect of recombinant EPO (rhEPO) in dystrophic muscles in mdx mice. Total strength was measured using a force transducer coupled to a computer. Gene expression for myostatin, transforming growth factor-β1 (TGF-β1), and tumor necrosis factor-α (TNF-α) was determined by quantitative real time polymerase chain reaction. Myostatin expression was significantly decreased in quadriceps from mdx mice treated with rhEPO (rhEPO = 0.60 ± 0.11, control = 1.07 ± 0.11). On the other hand, rhEPO had no significant effect on the expression of TGF-β1 (rhEPO = 0.95 ± 0.14, control = 1.05 ± 0.16) and TNF-α (rhEPO = 0.73 ± 0.20, control = 1.01 ± 0.09). These results may help to clarify some of the direct actions of EPO on skeletal muscle. PMID:25296358

  14. Familial polycythemia due to truncations of the erythropoietin receptor.

    PubMed Central

    Forget, B. G.; Degan, B. A.; Arcasoy, M. O.

    2000-01-01

    We studied a kindred with dominantly inherited familial erythrocytosis associated with heterozygosity for a deletion of seven nucleotides in exon 8 of the EpoR gene resulting in an EpoR peptide that is truncated by 59 amino acids in its C-terminal intracytoplasmic signal transduction domain. A seven basepair direct repeat sequence is present in the normal EpoR gene at the site of this mutation, consistent with the slipped mispairing model for the generation of short deletions during DNA replication. Hypersensitivity to erythropoietin of primary erythroid progenitors from an affected individual was observed in in vitro cultures of peripheral blood mononuclear cells, as indicated by the growth, at suboptimal concentrations of added Epo, of more numerous and larger BFU-E-derived erythroid cell colonies compared to those obtained from a normal control subject. To study mutant EpoR function, the cDNA encoding the mutant EpoR was stably transfected into murine growth factor (IL-3)-dependent 32D tissue culture cells. In proliferation assays, cells expressing the mutant EpoR displayed 5 to 10-fold increased sensitivity to Epo compared to cells expressing similar numbers of the wild type EpoR. In addition, the cells transfected with the mutant truncated receptor demonstrated prolonged activity of Jak2 kinase and Stat5 activity, molecules that mediate signal transduction by the EpoR. PMID:10881330

  15. Method of detecting genetically modified chicken containing human erythropoietin gene.

    PubMed

    Nakajima, Osamu; Nakamura, Kosuke; Kondo, Kazunari; Akiyama, Hiroshi; Teshima, Reiko

    2013-01-01

    Genetically modified (GM) chickens carrying the human erythropoietin (hEpo) gene have been developed to produce recombinant hEpo protein in eggs. However, such animals have not been approved as food sources in Japan. We developed a method for detecting the hEpo gene in chicken meat using a real-time polymerase chain reaction (real-time PCR). The hEpo gene was clearly detected in genomic DNA extracted from magnum and heart of a chimeric chicken containing the hEpo gene. A plasmid containing the hEpo gene was used as a standard reference molecule as well. The results clearly showed that our method was capable of detecting the hEpo gene contained in the plasmid in the presence of genomic DNA extracted from a raw chicken meat sample. We successfully used this method to test six samples of raw chicken meat and six samples of chicken meat in processed foods. This method will be useful for monitoring chicken meat that might have originated from GM chickens carrying the hEpo gene to assure food safety.

  16. The anemia of primary autonomic failure and its reversal with recombinant erythropoietin

    NASA Technical Reports Server (NTRS)

    Biaggioni, I.; Robertson, D.; Krantz, S.; Jones, M.; Haile, V.

    1994-01-01

    OBJECTIVE: To determine if chronic sympathetic deprivation is associated with anemia and a low erythropoietin response. DESIGN: Survey of the prevalence and characteristics of anemia in patients with severe primary autonomic failure. SETTING: A referral service for autonomic failure in a tertiary teaching hospital. PATIENTS: 84 patients with primary autonomic failure who had symptomatic orthostatic hypotension. INTERVENTION: Open-label trial with human recombinant erythropoietin. RESULTS: Anemia was present in 32 of 84 patients (38%; 95% Cl, 27% to 50%). Plasma norepinephrine levels, measured in patients standing upright, were lower in the patient group with lower hemoglobin levels. Mean values in 22 patients with a hemoglobin level of less than 120 g/L were as follows: hemoglobin, 108 g/L (range, 87 to 118 g/L); hematocrit, 0.33; corrected reticulocyte counts, 0.008; mean corpuscular volume, 89 fL (89 microns 3); serum iron, 16.5 mumol/L (92 micrograms/dL); total iron binding capacity, 43.3 mumol/L (242 micrograms/dL); ferritin, 184 micrograms/L; serum vitamin B12, 410 pmol/L (556 pg/mL); and serum folate, 22.7 nmol/L (10 ng/mL). No relation was found between serum erythropoietin and blood hemoglobin levels. In seven of nine patients with autonomic failure who had hemoglobin levels less than 120 g/L, serum erythropoietin levels decreased below the 95% confidence interval corresponding to patients with iron deficiency anemia. Therapy with recombinant erythropoietin improved mean hemoglobin levels (from 108 to 133 g/L) in all patients treated (n = 5) at relatively low doses (25 to 50 units/kg body weight, subcutaneously, three times a week). CONCLUSIONS: Our data support the hypothesis that the sympathetic nervous system stimulates erythropoiesis in humans because anemia is a frequent occurrence in patients with severe autonomic failure and is associated with a blunted erythropoietin response.

  17. Incidence of erythropoietin antibody-mediated pure red cell aplasia: the Prospective Immunogenicity Surveillance Registry (PRIMS)

    PubMed Central

    Macdougall, Iain C.; Casadevall, Nicole; Locatelli, Francesco; Combe, Christian; London, Gerard M.; Di Paolo, Salvatore; Kribben, Andreas; Fliser, Danilo; Messner, Hans; McNeil, John; Stevens, Paul; Santoro, Antonio; De Francisco, Angel L.M.; Percheson, Paul; Potamianou, Anna; Foucher, Arnaud; Fife, Daniel; Mérit, Véronique; Vercammen, Els

    2015-01-01

    Background Subcutaneous administration of Eprex® (epoetin alfa) in patients with chronic kidney disease (CKD) was contraindicated in the European Union between 2002 and 2006 after increased reports of anti-erythropoietin antibody-mediated pure red cell aplasia (PRCA). The Prospective Immunogenicity Surveillance Registry (PRIMS) was conducted to estimate the incidence of antibody-mediated PRCA with subcutaneous administration of a new coated-stopper syringe presentation of Eprex® and to compare this with the PRCA incidence with subcutaneous NeoRecormon® (epoetin beta) and Aranesp® (darbepoetin alfa). Methods PRIMS was a multicentre, multinational, non-interventional, parallel-group, immunogenicity surveillance registry. Adults with CKD receiving or about to initiate subcutaneous Eprex®, NeoRecormon® or Aranesp® for anaemia were enrolled and followed for up to 3 years. Unexplained loss or lack of effect (LOE), including suspected PRCA, was reported, with antibody testing for confirmation of PRCA. Results Of the 15 333 patients enrolled, 5948 received Eprex® (8377 patient-years) and 9356 received NeoRecormon®/Aranesp® (14 286 patient-years). No treatment data were available for 29 patients. Among 23 patients with LOE, five cases of PRCA were confirmed (Eprex®, n = 3; NeoRecormon®, n = 1; Aranesp®, n = 1). Based on exposed time, PRCA incidence was 35.8/100 000 patient-years (95% CI 7.4–104.7) for Eprex® versus 14.0/100 000 patient-years (95% CI 1.7–50.6) for NeoRecormon®/Aranesp®. The incidence of PRCA with Eprex® was not significantly different versus comparator ESAs (rate ratio: 2.56; 95% CI 0.43–15.31). An analysis based on observed time produced similar findings. Conclusion This large, prospective registry demonstrates that PRCA is rare with subcutaneous administration of either the new coated-stopper syringe presentation of Eprex®, or NeoRecormon® or Aranesp®. PMID:25239637

  18. Erythropoietin-induced acute erythroid leukemia-like picture: a potential pitfall.

    PubMed

    Moharram, Laila; Kamal, Nazmi; Al Sukhun, Sana; Sughayer, Maher A

    2014-03-01

    A 31-year-old male patient presented with fever and pancytopenia. He was diagnosed as a case of chronic anemia since early childhood. The etiology of the anemia was not known. The patient was transfusion dependent, and he had been maintained on erythropoietin for three years prior to admission. A bone marrow examination revealed prominent proliferation of immature and dysplastic erythroid precursors. Acute erythroid leukemia of the pure erythroid subtype was suspected. However, because of the history of erythropoietin therapy a definite diagnosis was not made. On follow-up one month later, the marrow changes had reversed to normal.

  19. Effect of recombinant human erythropoietin on mitomycin C-induced oxidative stress and genotoxicity in rat kidney and heart tissues.

    PubMed

    Rjiba-Touati, K; Ayed-Boussema, I; Guedri, Y; Achour, A; Bacha, H; Abid-Essefi, S

    2016-01-01

    Mitomycin C (MMC) is an antineoplastic agent used for the treatment of several human malignancies. Nevertheless, the prolonged use of the drug may result in a serious heart and kidney injuries. Recombinant human erythropoietin (rhEPO) has recently been shown to exert an important cytoprotective effect in experimental brain injury and ischemic acute renal failure. The aim of the present work is to investigate the cardioprotective and renoprotective effects of rhEPO against MMC-induced oxidative damage and genotoxicity. Our results showed that MMC induced oxidative stress and DNA damage. rhEPO administration in any treatment conditions decreased oxidative damage induced by MMC. It reduced malondialdehyde and protein carbonyl levels. rhEPO ameliorated reduced glutathione plus oxidized glutathione modulation and the increased catalase activity after MMC treatment. Furthermore, rhEPO restored DNA damage caused by MMC. We concluded that rhEPO administration especially in pretreatment condition protected rats against MMC-induced heart and renal oxidative stress and genotoxicity.

  20. A re-assessment of erythropoietin as a neuroprotective agent following rat spinal cord compression or contusion injury.

    PubMed

    Pinzon, Alberto; Marcillo, Alexander; Pabon, Diego; Bramlett, Helen M; Bunge, Mary Bartlett; Dietrich, W Dalton

    2008-09-01

    This study was initiated due to an NIH "Facilities of Research--Spinal Cord Injury" contract to support independent replication of published studies that appear promising for eventual clinical testing. We repeated a study reporting the beneficial effects of recombinant human erythropoietin (rhEPO) treatment after spinal cord injury (SCI). Moderate thoracic SCI was produced by two methods: 1) compression due to placement of a modified aneurysm clip (20 g, 10 s) at the T3 spinal segment (n=45) [followed by administration of rhEPO 1000 IU/kg/IP in 1 or 3 doses (treatment groups)] and 2) contusion by means of the MASCIS impactor (n = 42) at spinal T9 (height 12.5 cm, weight 10 g) [followed by the administration of rhEPO 5000 IU/kg/IP for 7d or single dose (treatment groups)]. The use of rhEPO following moderate compressive or contusive injury of the thoracic spinal cord did not improve the locomotor behavior (BBB rating scale). Also, secondary changes (i.e. necrotic changes followed by cavitation) were not significantly improved with rhEPO therapy. With these results, although we cannot conclude that there will be no beneficial effect in different SCI models, we caution researchers that the use of rhEPO requires further investigation before implementing clinical trials. PMID:18625498

  1. Erythropoietin messenger RNA levels in developing mice and transfer of /sup 125/I-erythropoietin by the placenta

    SciTech Connect

    Koury, M.J.; Bondurant, M.C.; Graber, S.E.; Sawyer, S.T.

    1988-07-01

    Erythropoietin (EP) mRNA was measured in normal and anemic mice during fetal and postnatal development. Normal fetal livers at 14 d of gestation contained a low level of EP mRNA. By day 19 of gestation, no EP mRNA was detected in normal or anemic fetal livers or normal fetal kidneys, but anemic fetal kidneys had low levels of EP mRNA. Newborn through adult stage mice responded to anemia by accumulating renal and hepatic EP mRNA. However, total liver EP mRNA was considerably less than that of the kidneys. Juvenile animals, 1-4 wk old, were hyperresponsive to anemia in that they produced more EP mRNA than adults. Moreover, nonanemic juveniles had readily measured renal EP mRNA, whereas the adult level was at the lower limit of detection. Because of the very low level of fetal EP mRNA, placental transfer of EP was evaluated. When administered to the pregnant mouse, /sup 125/I-EP was transferred in significant amounts to the fetuses. These results indicate that in mice the kidney is the main organ of EP production at all stages of postnatal development and that adult kidney may also play some role in providing EP for fetal erythropoiesis via placental transfer of maternal hormone.

  2. Erythropoietin in heart failure and other cardiovascular diseases: hematopoietic and pleiotropic effects.

    PubMed

    Manolis, Antonis S; Tzeis, Stylianos; Triantafyllou, Kostas; Michaelidis, John; Pyrros, Ioannis; Sakellaris, Nikolaos; Kranidis, Athanasios; Melita, Helen

    2005-10-01

    Erythropoietin is a hypoxia-induced hormone that is a major regulator of normal erythropoiesis. Over the last decade, the production of recombinant human erythropoietin has revolutionized the treatment of anemia associated with chronic renal failure, and has led to a greater understanding of anemia pathophysiology and to the elucidation of the interactions of erythropoietin, iron, and erythropoiesis. Anemia has been shown to be independently associated with increased mortality and disease progression. Potential survival benefits associated with correction of anemia have expanded considerably the indications of erythropoietin use in various patient populations and are leading to consideration of earlier, more aggressive treatment of mild to moderate anemia. The results of such treatment are promising in a variety of new clinical settings, including anemia associated with congestive heart failure. Furthermore, the erythropoietin receptor is widely distributed in the cardiovascular system, including endothelial cells, smooth muscle cells and cardiomyocytes and preclinical studies have established erythropoietin to be a pleiotropic cytokine with anti-apoptotic activity and tissue-protective actions in the cardiovascular system, beyond correction of hemoglobin levels. Despite some potential adverse effects, such as hypertension, and the occurrence of erythropoietin resistance, early studies in heart failure patients with anemia suggest that erythropoietin therapy is safe and effective in reducing left ventricular hypertrophy, enhancing exercise performance and increasing ejection fraction. Anemia is found in about one-third of all cases of congestive heart failure (CHF). The most likely common cause is chronic renal insufficiency, which is present in about half of all CHF cases. However, anemia can occur in CHF without renal insufficiency and is likely to be due to excessive cytokine production. The anemia itself can worsen cardiac function, both because it causes

  3. Darbepoietin-alfa has comparable erythropoietic stimulatory effects to recombinant erythropoietin whilst preserving the bone marrow microenvironment

    PubMed Central

    Dewamitta, Sita R.; Russell, Megan R.; Nandurkar, Harshal; Walkley, Carl R.

    2013-01-01

    Erythropoiesis stimulating agents are widely used for the treatment of anemia. Recently, we reported erythroid expansion with impaired B lymphopoiesis and loss of trabecular bone in C57BL/6 mice following ten days of treatment with low-dose short acting recombinant human erythropoietin. We have assessed erythropoietin against longer-acting darbepoietin-alfa at a comparable erythroid stimulatory dosage regime. Darbepoietin-alfa and erythropoietin induced similar in vivo erythropoietic expansion. Both agents induced an expansion of the colony-forming unit-erythroid populations. However, unlike erythropoietin, darbepoietin-alfa did not impair bone marrow B lymphopoiesis. Strikingly the bone loss observed with erythropoietin was not apparent following darbepoietin-alfa treatment. This analysis demonstrates that whilst darbepoietin-alfa has similar in vivo erythropoietic potency to erythropoietin, it preserves the bone marrow microenvironment. Thus erythropoietin and darbepoietin-alfa manifest different action showing that erythropoiesis stimulating agents have differential non-erythroid effects dependent on their duration of action. PMID:23242598

  4. Charting a course for erythropoietin in traumatic brain injury

    PubMed Central

    Maiese, Kenneth

    2016-01-01

    Traumatic brain injury (TBI) is a severe public health problem that impacts more than four million individuals in the United States alone and is increasing in incidence on a global scale. Importantly, TBI can result in acute as well as chronic impairments for the nervous system leaving individuals with chronic disability and in instances of severe trauma, death becomes the ultimate outcome. In light of the significant negative health consequences of TBI, multiple therapeutic strategies are under investigation, but those focusing upon the cytokine and growth factor erythropoietin (EPO) have generated a great degree of enthusiasm. EPO can control cell death pathways tied to apoptosis and autophagy as well oversees processes that affect cellular longevity and aging. In vitro studies and experimental animal models of TBI have shown that EPO can restore axonal integrity, promote cellular proliferation, reduce brain edema, and preserve cellular energy homeostasis and mitochondrial function. Clinical studies for neurodegenerative disorders that involve loss of cognition or developmental brain injury support a positive role for EPO to prevent or reduce injury in the nervous system. However, recent clinical trials with EPO and TBI have not produced such clear conclusions. Further clinical studies are warranted to address the potential efficacy of EPO during TBI, the concerns with the onset, extent, and duration of EPO therapeutic strategies, and to focus upon the specific downstream pathways controlled by EPO such as protein kinase B (Akt), mechanistic target of rapamycin (mTOR), AMP activated protein kinase (AMPK), sirtuins, wingless pathways, and forkhead transcription factors for improved precision against the detrimental effects of TBI. PMID:27081573

  5. Immunosuppressive Effects of Erythropoietin on Human Alloreactive T Cells

    PubMed Central

    Cravedi, Paolo; Manrique, Joaquin; Hanlon, Katherine E.; Reid-Adam, Jessica; Brody, Joshua; Prathuangsuk, Praeophayom; Mehrotra, Anita

    2014-01-01

    Correction of anemia with erythropoietin (EPO) is associated with improved kidney transplant outcomes. Emerging evidence, predominantly from animal models, indicates that these observations may be erythropoiesis-independent and that EPO exhibits immunosuppressive properties. We examined the effects of EPO on human T-cell alloimmunity by first documenting that CD4+ and CD8+ T cells express EPO receptor (EPO-R) on their surfaces. In mixed lymphocyte reactions, EPO induced a dose-dependent decrease in allogeneic CD4+ T-cell proliferation (EPO 1000 U/ml: 44.6%±22.9% of vehicle, P<0.05; 2000 U/ml: 11.1%±4% of vehicle, P<0.001) without inducing cell death. The effects required signals transmitted directly through the EPO-R expressed on T cells, resulting in diminished Th1 differentiation without effects on regulatory T-cell induction. Mechanistic studies revealed that EPO prevented IL-2–induced proliferation by uncoupling IL-2 receptor signaling, inhibiting phosphorylation of the intracellular intermediaries AKT and extracellular signal-regulated kinase that are known to mediate T-cell expansion. EPO treatment reduced expansion of human naïve CD4+ T cells after adoptive transfer into NOD scid γcnull mouse recipients, verifying the effects in vivo. Although activated T cells expressed CD131, an alternative EPO receptor, addition of a specific CD131 agonist peptide, ARA290, did not alter T-cell proliferation or cytokine production. Our findings link EPO-R signaling on T cells to inhibition of T-cell immunity, providing one mechanism that could explain the observed protective effects of EPO in kidney transplant recipients. PMID:24676641

  6. Cost-effectiveness of continuous erythropoietin receptor activator in anemia

    PubMed Central

    Schmid, Holger

    2014-01-01

    Background Erythropoiesis-stimulating agents (ESAs) are the mainstay of anemia therapy. Continuous erythropoietin receptor activator (CERA) is a highly effective, long-acting ESA developed for once-monthly dosing. A multitude of clinical studies has evaluated the safety and efficiency of this treatment option for patients with renal anemia. In times of permanent financial pressure on health care systems, the cost-effectiveness of CERA should be of particular importance for payers and clinicians. Objective To critically analyze, from the nephrologists’ point of view, the published literature focusing on the cost-effectiveness of CERA for anemia treatment. Methods The detailed literature search covered electronic databases including MEDLINE, PubMed, and Embase, as well as international conference abstract databases. Results Peer-reviewed literature analyzing the definite cost-effectiveness of CERA is scarce, and most of the available data originate from conference abstracts. Identified data are restricted to the treatment of anemia due to chronic kidney disease. Although the majority of studies suggest a considerable cost advantage for CERA, the published literature cannot easily be compared. While time and motion studies clearly indicate that a switch to CERA could minimize health care staff time in dialysis units, the results of studies comparing direct costs are more ambivalent, potentially reflecting the differences between health care systems and variability between centers. Conclusion Analyzed data are predominantly insufficient; they miss clear evidence and have to thus be interpreted with great caution. In this day and age of financial restraints, results from well-designed, head-to-head studies with clearly defined endpoints have to prove whether CERA therapy can achieve cost savings without compromising anemia management. PMID:25050070

  7. [Erythropoietin and drug resistance in breast and ovarian cancers].

    PubMed

    Szenajch, Jolanta M; Synowiec, Agnieszka E

    2016-01-01

    Recombinant human erythropoietin (rhEPO) is used in breast and ovarian cancer patients to alleviate cancer- and chemotherapy-related anemia. Some clinical trials have reported that rhEPO may adversely impact survival and increase the risk of thrombovascular events in patients with breast cancer but not with ovarian cancer. The latter may potentially benefit the most from rhEPO treatment due to the nephrotoxic and myelosuppresive effects of standard platinum-based chemotherapy used in ovarian cancer disease. However, over the last decade the preclinical data have revealed that EPO is not only the principal growth factor and the hormone which regulates erythropoiesis, but also a cytokine with a pleiotropic activity which also can affect cancer cells. EPO can stimulate survival, ability to form metastases and drug resistance not only in continuous breast- and ovarian cancer cell lines but also in breast cancer stem-like cells. EPO receptor (EPOR) can also be constitutively active in both these cancers and, in breast cancer cells, may act in an interaction with estrogen receptor (ER) and epidermal growth factor receptor-2 (HER-2). EPOR, by an EPO-independent mechanism, promotes proliferation of breast cancer cells in cooperation with estrogen receptor, resulting in decreased effectiveness of tamoxifen treatment. In another interaction, as a result of the molecular antagonism between EPOR and HER2, rhEPO protects breast cancer cells against trastuzumab. Both clinical and preclinical evidence strongly suggest the urgent need to reevaluate the traditional use of rhEPO in the oncology setting.

  8. Erythropoietin Augments Survival of Glioma Cells After Radiation and Temozolomide

    SciTech Connect

    Hassouna, Imam; Sperling, Swetlana; Kim, Ella; Schulz-Schaeffer, Walter; Rave-Fraenk, Margret; Hasselblatt, Martin; Jelkmann, Wolfgang; Giese, Alf; Ehrenreich, Hannelore

    2008-11-01

    Purpose: Despite beneficial effects of irradiation/chemotherapy on survival of glioblastoma (GBM) patients, collateral damage to intact neural tissue leads to 'radiochemobrain' and reduced quality of life in survivors. For prophylactic neuroprotection, erythropoietin (EPO) is a promising candidate, provided that concerns regarding potential tumor promoting effects are alleviated. Methods and Materials: Human GBM-derived cell lines U87, G44, G112, and the gliosarcoma-derived line G28 were treated with EPO, with and without combinations of irradiation or temozolomide (TMZ). Responsiveness of glioma cells to EPO was measured by cell migration from spheroids, cell proliferation, and clonogenic survival. Implantation of U87 cells into brains of nude mice, followed 5 days later by EPO treatment (5,000 U/kg intraperitoneal every other day for 2 weeks) should reveal effects of EPO on tumor growth in vivo. Reverse transcriptase-polymerase chain reaction was performed for EPOR, HIF-1{alpha}, and epidermal growth factor receptor (EGFR)vIII in cell lines and 22 human GBM specimens. Results: EPO did not modulate basal glioma cell migration and stimulated proliferation in only one of four cell lines. Importantly, EPO did not enhance tumor growth in mouse brains. Preincubation of glioma cells with EPO for 3 h, followed by irradiation and TMZ for another 24 h, resulted in protection against chemoradiation-induced cytotoxicity in three cell lines. Conversely, EPO induced a dose-dependent decrease in survival of G28 gliosarcoma cells. In GBM specimens, expression of HIF-1{alpha} correlated positively with expression of EPOR and EGFRvIII. EPOR and EGFRvIII expression did not correlate. Conclusions: EPO is unlikely to appreciably influence basal glioma growth. However, concomitant use of EPO with irradiation/chemotherapy in GBM patients is not advisable.

  9. Erythropoietin modulates intracellular calcium in a human neuroblastoma cell line

    PubMed Central

    Assandri, Roberta; Egger, Marcel; Gassmann, Max; Niggli, Ernst; Bauer, Christian; Forster, Ian; Görlach, Agnes

    1999-01-01

    Recent investigations have shown that the glycoprotein erythropoietin (Epo) and its specific receptor (EpoR) are present in the mammalian brain including human, monkey and mouse. These findings suggest a local action of Epo in the nervous system. The aim of this study was to elucidate a possible functional interaction of Epo with neuronal cells. To examine the influence of externally applied Epo on Ca2+ homeostasis the human neuroblastoma cell line SK-N-MC was chosen as a suitable in vitro model for undifferentiated neuronal cells. Expression of the EpoR in SK-N-MC cells was detected by reverse transcription-PCR, Western blot and immunofluorescence analysis. Patch-clamp studies of SK-N-MC cells confirmed the expression of T-type Ca2+ channels, whose peak macroscopic current was increased by the addition of recombinant human Epo (rhEpo) to the bathing medium. Confocal laser scanning microscopy analysis of SK-N-MC cells confirmed a transient increase in intracellular free [Ca2+] in response to externally applied rhEpo. The transient response to Epo was dependent on external Ca2+ and remained even after depletion of internal Ca2+ stores by caffeine or thapsigargin. However, after depletion the response to Epo was absent when cells were superfused with the T-type Ca2+ channel blocker flunarizine. This study demonstrates that Epo can interact with neuronal cells by affecting Ca2+ homeostasis through an increase in Ca2+ influx via plasma membrane T-type voltage-dependent Ca2+ channels. PMID:10087335

  10. Immunosuppressive effects of erythropoietin on human alloreactive T cells.

    PubMed

    Cravedi, Paolo; Manrique, Joaquin; Hanlon, Katherine E; Reid-Adam, Jessica; Brody, Joshua; Prathuangsuk, Praeophayom; Mehrotra, Anita; Heeger, Peter S

    2014-09-01

    Correction of anemia with erythropoietin (EPO) is associated with improved kidney transplant outcomes. Emerging evidence, predominantly from animal models, indicates that these observations may be erythropoiesis-independent and that EPO exhibits immunosuppressive properties. We examined the effects of EPO on human T-cell alloimmunity by first documenting that CD4(+) and CD8(+) T cells express EPO receptor (EPO-R) on their surfaces. In mixed lymphocyte reactions, EPO induced a dose-dependent decrease in allogeneic CD4(+) T-cell proliferation (EPO 1000 U/ml: 44.6%±22.9% of vehicle, P<0.05; 2000 U/ml: 11.1%±4% of vehicle, P<0.001) without inducing cell death. The effects required signals transmitted directly through the EPO-R expressed on T cells, resulting in diminished Th1 differentiation without effects on regulatory T-cell induction. Mechanistic studies revealed that EPO prevented IL-2-induced proliferation by uncoupling IL-2 receptor signaling, inhibiting phosphorylation of the intracellular intermediaries AKT and extracellular signal-regulated kinase that are known to mediate T-cell expansion. EPO treatment reduced expansion of human naïve CD4(+) T cells after adoptive transfer into NOD scid γc(null) mouse recipients, verifying the effects in vivo. Although activated T cells expressed CD131, an alternative EPO receptor, addition of a specific CD131 agonist peptide, ARA290, did not alter T-cell proliferation or cytokine production. Our findings link EPO-R signaling on T cells to inhibition of T-cell immunity, providing one mechanism that could explain the observed protective effects of EPO in kidney transplant recipients.

  11. Control of erythropoietin gene expression and its use in medicine.

    PubMed

    Jelkmann, Wolfgang

    2007-01-01

    Erythropoietin (EPO) gene expression is under the control of inhibitory (GATA-2, NF-kappaB) and stimulatory (hypoxia-inducible transcription factor [HIF]-2, hepatocyte nuclear factor [HNF]-4alpha [alpha]) transcription factors. EPO deficiency is the main cause of the anemia in chronic kidney disease (CKD) and a contributing factor in the anemias of inflammation and cancer. Small, orally active compounds capable of stimulating endogenous EPO production are in preclinical or clinical trials for treatment of anemia. These agents include stabilizers of the HIFs that bind to the EPO enhancer and GATA inhibitors which prevent GATA from suppressing the EPO promoter. While HIF stabilizing drugs may prove useful as inexpensive second-line choices, at present, their side effects--particularly tumorigenicity--preclude their use as first-choice therapy. As an alternative, EPO gene therapy has been explored in animal studies and in trials on CKD patients. Here, a major problem is immunogenicity of ex vivo transfected implanted cells and of the recombinant protein produced after ex vivo or in vivo EPO complementary DNA (cDNA) transfer. Recombinant human EPO (rhEPO) engineered in Chinese hamster ovary (CHO) cell cultures (epoetin alpha and epoetin beta [beta]) and its hyperglycosylated analogue darbepoetin alpha are established and safe drugs to avoid allogeneic red blood cell transfusion. Gene-activated EPO (epoetin delta [delta]) from human fibrosarcoma cells (HT-1080) has recently been launched for use in CKD. It is important to know the basics of the technologies, production processes, and structural properties of the novel anti-anemic strategies and drugs. PMID:17998055

  12. Pharmacokinetic and pharmacodynamic studies following percutaneous absorption of erythropoietin micropiles to rats.

    PubMed

    Ito, Y; Shiroyama, K; Yoshimitsu, J; Ohashi, Y; Sugioka, N; Takada, K

    2007-08-28

    To ascertain the pharmacological activity of erythropoietin (EPO) administered by self-dissolving micropiles (SDMP), four kinds of EPO SDMPs were prepared and were administered to rats in 4 consecutive days at 200, 500, 1000 and 2300 IU/kg. After the start of the experiment, blood samples were obtained once a day for 10 days and percent circulating reticulocytes were counted using Miller technique. At the lower doses, 200 and 500 IU/kg, pharmacological activity of EPO was not obtained. By increasing EPO dose to 1000 IU/kg, circulating reticulocytes significantly increased at days 4, 5, 6 and 7 after the start of the experiment and the average value for the change in reticulocyte levels during day 1 and day 5 was 170.9%. With the highest dose, 2300 IU/kg, higher circulating reticulocytes levels started to increase at the 4th day after the start of the experiment and maintained from day 5 to day 10. The average of the changes in reticulocyte from day 5 to day 10 was 251%. Dose-dependent circulating reticulocytes increase was observed at the higher dose range, 1000 and 2300 IU/kg. To study the linearity on the serum EPO level vs. time curves, pharmacokinetic experiment was performed with rats. After the administration of EPO SDMPs to rats, 200, 500, 1000 and 2300 IU/kg, serum EPO levels gradually increased and reached to the maximum level, C(max), at 18 h after administration. The C(max)s were 100.4+/-11.7 mIU/ml (200 IU/kg), 346.6+/-11.8 mIU/ml (500 IU/kg), 391.6+/-17.6 mIU/ml (1000 IU/kg), and 1094.9+/-114.8 mIU/ml (2300 IU/kg), respectively. AUCs were 1407+/-231, 3843+/-402, 5363+/-482 and 15,566+/-1894 mIU h/ml. Linear relation was obtained between serum EPO level and EPO dose administered as SDMP. With histological study, any adverse effect was not found out on the skin where SDMPs were administered for consecutive 4 days. These results suggest the usefulness of SDMP as a new percutaneous delivery system of EPO.

  13. Disease Activity and Conversion into Multiple Sclerosis after Optic Neuritis Is Treated with Erythropoietin

    PubMed Central

    Sühs, Kurt-Wolfram; Papanagiotou, Panagiotis; Hein, Katharina; Pul, Refik; Scholz, Kerstin; Heesen, Christoph; Diem, Ricarda

    2016-01-01

    Changes in cerebral lesion load by magnetic resonance imaging (MRI) in patients from a double-blind, placebo-controlled, phase II study on erythropoietin in clinically isolated optic neuritis (ClinicalTrials.gov, NCT00355095) were analyzed. Therefore, patients with acute optic neuritis were assigned to receive either 33,000 IU of recombinant human erythropoietin (IV) daily for three days, or a placebo, as an add-on to methylprednisolone. Of 35 patients, we investigated changes in cerebral lesion load in MRIs obtained at baseline and at weeks 4, 8, and 16. In 5 of the 35 patients, we found conversion into multiple sclerosis (MS) based on MRI progression only. These five patients had received the placebo. Another five patients showed MRI progression together with relapses. Three of these patients had received erythropoietin, and two the placebo. Yet, analyzing the change in absolute numbers of periventricular, juxtacortical, and infratentorial lesions including gadolinium-enhancing lesions, there were no significant differences between the groups. Although effective in terms of retinal nerve fiber layer protection, erythropoietin treatment of acute isolated optic neuritis did not influence further evolution of MRI lesions in the brain when comparing absolute numbers. However, early conversion from clinically isolated syndrome to MS assessed by MRI activity seemed to occur more frequently in the placebo-treated group. PMID:27706045

  14. Androgen versus erythropoietin for the treatment of anaemia of pre-dialysis chronic kidney disease.

    PubMed

    Paul, A K; Latif, Z A; Iqbal, S; Amin, F; Shefin, S M; Ashrafuzzaman, S M

    2012-01-01

    Chronic kidney disease is a microvascular complication of diabetes mellitus (DM). Anemia is an important clinical manifestation to treat chronic kidney disease. Many subjects with poor socio-economic status having chronic kidney disease (CKD) and anaemia in a developing country can not afford the treatment with erythropoietin. This study has designed to see the efficacy of Nandrolone, a cheaper alternative; in comparison with recombinant human erythropoietin for management of anemia of pre-dialysis diabetic chronic kidney disease. Sixty adult diabetic patients with anaemia of chronic kidney disease on conservative treatment [Not on Hemodialysis (HD)] were enrolled. Patients were divided into two groups (Group 1 and Group 2) of 30 patients each. Group 1 patients received nandrolone deaconate 50 mg deep intramuscular and Group 2 recombinant human erythropoietin 100 IU per kilogram of body weight subcutaneously once weekly. Patients of both group received oral supplements in order to maintain body iron stores. All the relevant haematological and renal parameters were evaluated at the end of 3rd & 6th months. There was a statistically significant rise in haemoglobin concentration, packed cell volume, in both groups. The rise in haemoglobin concentration, in Group 2 was more marked followed by Group 1, at the end of 3rd, and 6th months. Nandrolone, though not equally effective, may be considered as a valid alternative therapy for the treatment of anemia of pre-dialysis diabetic chronic kidney disease to that of erythropoietin.

  15. Enhanced brain release of erythropoietin, cytokines and NO during carotid clamping.

    PubMed

    Carelli, Stephana; Ghilardi, Giorgio; Bianciardi, Paola; Latorre, Elisa; Rubino, Federico; Bissi, Marina; Di Giulio, Anna Maria; Samaja, Michele; Gorio, Alfredo

    2016-02-01

    Although effective and safe, carotid endarterectomy (CEA) implies a reduced blood flow to the brain and likely an ischemia/reperfusion event. The high rate of uneventful outcomes associated with CEA suggests the activation of brain endogenous protection mechanisms aimed at limiting the possible ischemia/reperfusion damage. This study aims at assessing whether CEA triggers protective mechanisms such as brain release of erythropoietin and nitric oxide. CEA was performed in 12 patients; blood samples were withdrawn simultaneously from the surgically exposed ipsilateral jugular and leg veins before, during (2 and 40 min) and after clamp removal (2 min). Plasma antioxidant capacity, carbonylated proteins, erythropoietin, nitrates and nitrites (NOx) were determined. No changes in intraoperative EEG, peripheral and transcranial blood oxygen saturation were detectable, and no patients showed any neurologic sign after the intervention. Antioxidant capacity and protein carbonylation in plasma were unaffected. Differently, erythropoietin, VEGF, TNF-α and NOx increased during clamping in the jugular blood (2 and 40 min), while no changes were observed in the peripheral circulation. These results show that blood erythropoietin, VEGF, TNF-α, and NOx increased in the brain during uncomplicated CEA. This may represent an endogenous self-activated neuroprotective mechanism aimed at the prevention of ischemia/reperfusion damage. PMID:26494654

  16. The Winding Pathway to Erythropoietin Along the Chemistry–Biology Frontier: A Success At Last

    PubMed Central

    Wilson, Rebecca M.; Dong, Suwei; Wang, Ping; Danishefsky, Samuel J.

    2016-01-01

    The total synthesis of a homogeneous erythropoietin, possessing the native amino acid sequence and chitobiose glycans at each of the three wild-type sites of N-glycosylation, has been accomplished in our laboratory. We provide herein an account of our decade-long research effort en route to this formidable target compound. PMID:23775885

  17. Loss of Quiescence in von Hippel-Lindau Hemangioblastomas is Associated with Erythropoietin Signaling

    PubMed Central

    Feldman, Michael J.; Sizdahkhani, Saman; Edwards, Nancy A.; Merrill, Marsha J.; Ray-Chaudhury, Abhik; Zhuang, Zhengping; Lonser, Russell R.; Oldfield, Edward H.; Chittiboina, Prashant

    2016-01-01

    von Hippel-Lindau (VHL) patients develop multiple central nervous system hemangioblastomas (HB). Some HBs become symptomatic with exponential growth or cyst formation following long periods of quiescence. Understanding the factors underlying growth in hemangioblastoma may lead to better strategies to arrest or prevent tumor growth. In 5 VHL patients, we resected quiescent hemangioblastomas (Q-HB) that were en-route during surgical access to symptomatic hemangioblastomas (S-HB), for matched tumor analysis. Quantitative reverse transcriptase analysis demonstrated a 2-fold increase in EPO expression in all S-HB, while 4/5 showed either Hypoxia Inducible Factor-1α or 2α upregulation. Additionally, all S-HB had increased phosphorylated erythropoietin (EPO) receptor and phosphorylated STAT-5 relative to matched Q-HB, with increased phosphorylated JAK-2 largely confined to the stromal cells in clusters within the tumors. These findings suggest that Q-HB to S-HB conversion may be associated with an erythropoietin-signaling loop. Furthermore, we found that EPO is detectable in cyst fluid from S-HB (n = 14), while absent in CSF (n = 1). Additionally, S-HB presentation or S-HB resection does not result in discernible change in serum EPO or hemoglobin (n = 60). These observations suggest that the altered erythropoietin signaling is focal and suggests that studying modulation of erythropoietin receptor pathway may lead to strategies in preventing HB growth. PMID:27748427

  18. Erythropoietin stimulation of human adipose tissue for therapeutic refilling releases protective cytokines

    PubMed Central

    Sabbatini, Maurizio; Bosetti, Michela; Borrone, Alessia; Moalem, Liah; Taveggia, Antonio; Verna, Giovanni; Cannas, Mario

    2016-01-01

    Apoptosis and inflammatory processes may be at the basis of reducing graft survival. Erythropoietin is a tissue-protective hormone with pleiotropic potential, and it interferes with the activities of pro-inflammatory cytokines and stimulates healing following injury, preventing destruction of tissue surrounding the injury site. It may represent a useful tool to increase the autograft integration. Through the use of multipanel kit cytokine analysis we have detected the cytokines secreted by human tissue adipose mass seeded in culture following withdrawal by Coleman’s modified technique in three groups: control, after lipopolysaccharides stimulation and after erythropoietin stimulation. In the control group, we have observed expression of factors that may have a role in protecting the tissue homeostatic mechanism. But the same factors were secreted following stimulation with lipopolysaccharides combined with others factors that delineated the inflammatory state. Instead through erythropoietin stimulation, the factors known to exert tissue-protective action were secreted. Therefore, the use of a trophic factors such as erythropoietin may help to inhibit the potential inflammatory process development and stimulate the activation of reparative/regenerative process in the tissue graft. PMID:27738510

  19. Computational and functional analysis of biopharmaceutical drugs in zebrafish: Erythropoietin as a test model.

    PubMed

    Guarienti, Michela; Giacopuzzi, Edoardo; Gianoncelli, Alessandra; Sigala, Sandra; Spano, Pierfranco; Pecorelli, Sergio; Pani, Luca; Memo, Maurizio

    2015-12-01

    The zebrafish (Danio rerio) is a very popular vertebrate model system, especially embryos represent a valuable tool for in vivo pharmacological assays. This is mainly due to the zebrafish advantages when compared to other animal models. Erythropoietin is a glycoprotein hormone that acts principally on erythroid progenitors, stimulating their survival, proliferation and differentiation. Recombinant human erythropoietin (rhEPO) has been widely used in medicine to treat anemia and it is one of the best-selling biotherapeutics worldwide. The recombinant molecule, industrially produced in CHO cells, has the same amino acid sequence of endogenous human erythropoietin, but differs in the glycosylation pattern. This may influence efficacy and safety, particularly immunogenicity, of the final product. We employed the zebrafish embryo as a vertebrate animal model to perform in vivo pharmacological assays. We conducted a functional analysis of rhEPO alpha Eprex(®) and two biosimilars, the erythropoietin alpha Binocrit(®) and zeta Retacrit(®). By in silico analysis and 3D modeling we proved the interaction between recombinant human erythropoietin and zebrafish endogenous erythropoietin receptor. Then we treated zebrafish embryos with the 3 rhEPOs and we investigated their effect on erythrocytes production with different assays. By real time-PCR we observed the relative upregulation of gata1 (2.4 ± 0.3 fold), embryonic α-Hb (1.9 ± 0.2 fold) and β-Hb (1.6 ± 0.1 fold) transcripts. A significant increase in Stat5 phosphorylation was also assessed in embryos treated with rhEPOs when compared with the negative controls. Live imaging in tg (kdrl:EGFP; gata1:ds-red) embryos, o-dianisidine positive area quantification and cyanomethemoglobin content quantification revealed a 1.8 ± 0.3 fold increase of erythrocytes amount in embryos treated with rhEPOs when compared with the negative controls. Finally, we verified that recombinant human erythropoietins did not cause any

  20. Erythropoietin Treatment in Traumatic Brain Injury: Operation Brain Trauma Therapy.

    PubMed

    Bramlett, Helen M; Dietrich, W Dalton; Dixon, C Edward; Shear, Deborah A; Schmid, Kara E; Mondello, Stefania; Wang, Kevin K W; Hayes, Ronald L; Povlishock, John T; Tortella, Frank C; Kochanek, Patrick M

    2016-03-15

    Experimental studies targeting traumatic brain injury (TBI) have reported that erythropoietin (EPO) is an endogenous neuroprotectant in multiple models. In addition to its neuroprotective effects, it has also been shown to enhance reparative processes including angiogenesis and neurogenesis. Based on compelling pre-clinical data, EPO was tested by the Operation Brain Trauma Therapy (OBTT) consortium to evaluate therapeutic potential in multiple TBI models along with biomarker assessments. Based on the pre-clinical TBI literature, two doses of EPO (5000 and 10,000 IU/kg) were tested given at 15 min after moderate fluid percussion brain injury (FPI), controlled cortical impact (CCI), or penetrating ballistic-like brain injury (PBBI) with subsequent behavioral, histopathological, and biomarker outcome assessments. There was a significant benefit on beam walk with the 5000 IU dose in CCI, but no benefit on any other motor task across models in OBTT. Also, no benefit of EPO treatment across the three TBI models was noted using the Morris water maze to assess cognitive deficits. Lesion volume analysis showed no treatment effects after either FPI or CCI; however, with the 5000 IU/kg dose of EPO, a paradoxical increase in lesion volume and percent hemispheric tissue loss was seen after PBBI. Biomarker assessments included measurements of glial fibrillary acidic protein (GFAP) and ubiquitin C-terminal hydrolase-L1 (UCH-L1) in blood at 4 or 24 h after injury. No treatment effects were seen on biomarker levels after FPI, whereas treatment at either dose exacerbated the increase in GFAP at 24 h in PBBI but attenuated 24-4 h delta UCH-L1 levels at high dose in CCI. Our data indicate a surprising lack of efficacy of EPO across three established TBI models in terms of behavioral, histopathological, and biomarker assessments. Although we cannot rule out the possibility that other doses or more prolonged treatment could show different effects, the lack of efficacy of EPO reduced

  1. Erythropoietin Treatment in Traumatic Brain Injury: Operation Brain Trauma Therapy.

    PubMed

    Bramlett, Helen M; Dietrich, W Dalton; Dixon, C Edward; Shear, Deborah A; Schmid, Kara E; Mondello, Stefania; Wang, Kevin K W; Hayes, Ronald L; Povlishock, John T; Tortella, Frank C; Kochanek, Patrick M

    2016-03-15

    Experimental studies targeting traumatic brain injury (TBI) have reported that erythropoietin (EPO) is an endogenous neuroprotectant in multiple models. In addition to its neuroprotective effects, it has also been shown to enhance reparative processes including angiogenesis and neurogenesis. Based on compelling pre-clinical data, EPO was tested by the Operation Brain Trauma Therapy (OBTT) consortium to evaluate therapeutic potential in multiple TBI models along with biomarker assessments. Based on the pre-clinical TBI literature, two doses of EPO (5000 and 10,000 IU/kg) were tested given at 15 min after moderate fluid percussion brain injury (FPI), controlled cortical impact (CCI), or penetrating ballistic-like brain injury (PBBI) with subsequent behavioral, histopathological, and biomarker outcome assessments. There was a significant benefit on beam walk with the 5000 IU dose in CCI, but no benefit on any other motor task across models in OBTT. Also, no benefit of EPO treatment across the three TBI models was noted using the Morris water maze to assess cognitive deficits. Lesion volume analysis showed no treatment effects after either FPI or CCI; however, with the 5000 IU/kg dose of EPO, a paradoxical increase in lesion volume and percent hemispheric tissue loss was seen after PBBI. Biomarker assessments included measurements of glial fibrillary acidic protein (GFAP) and ubiquitin C-terminal hydrolase-L1 (UCH-L1) in blood at 4 or 24 h after injury. No treatment effects were seen on biomarker levels after FPI, whereas treatment at either dose exacerbated the increase in GFAP at 24 h in PBBI but attenuated 24-4 h delta UCH-L1 levels at high dose in CCI. Our data indicate a surprising lack of efficacy of EPO across three established TBI models in terms of behavioral, histopathological, and biomarker assessments. Although we cannot rule out the possibility that other doses or more prolonged treatment could show different effects, the lack of efficacy of EPO reduced

  2. Serum erythropoietin and outcome after ischaemic stroke: a prospective study

    PubMed Central

    Åberg, N David; Stanne, Tara M; Jood, Katarina; Schiöler, Linus; Blomstrand, Christian; Andreasson, Ulf; Blennow, Kaj; Zetterberg, Henrik; Isgaard, Jörgen; Jern, Christina; Svensson, Johan

    2016-01-01

    Objectives Erythropoietin (EPO), which is inversely associated with blood haemoglobin (Hb), exerts neuroprotective effects in experimental ischaemic stroke (IS). However, clinical treatment trials have so far been negative. Here, in patients with IS, we analysed whether serum EPO is associated with (1) initial stroke severity, (2) recovery and (3) functional outcome. Design Prospective. Controls available at baseline. Setting A Swedish hospital-initiated study with outpatient follow-up after 3 months. Participants Patients (n=600; 64% males, mean age 56 years, controls n=600) were included from the Sahlgrenska Academy Study on IS (SAHLSIS). Primary and secondary outcome measures In addition to EPO and Hb, initial stroke severity was assessed by the Scandinavian Stroke Scale (SSS) and compared with SSS after 3 months (follow-up) as a measure of recovery. Functional outcome was evaluated using the modified Rankin Scale (mRS) at follow-up. Serum EPO and SSS were divided into quintiles in the multivariate regression analyses. Results Serum EPO was 21% and 31% higher than in controls at the acute phase of IS and follow-up, respectively. In patients, acute serum EPO was 19.5% higher in severe versus mild IS. The highest acute EPO quintile adjusted for sex, age and Hb was associated with worse stroke severity quintile (OR 1.70, 95% CI 1.00 to 2.87), better stroke recovery quintile (OR 1.93, CI 1.09 to 3.41) and unfavourable mRS 3–6 (OR 2.59, CI 1.15 to 5.80). However, the fourth quintile of EPO increase (from acute to follow-up) was associated with favourable mRS 0–2 (OR 3.42, CI 1.46 to 8.03). Only the last association withstood full adjustment. Conclusions The crude associations between EPO and worse stroke severity and outcome lost significance after multivariate modelling. However, in patients in whom EPO increased, the association with favourable outcome remained after adjustment for multiple covariates. PMID:26916692

  3. Design and rationale of the Reduction of Infarct Expansion and Ventricular Remodeling with Erythropoietin After Large Myocardial Infarction (REVEAL) trial

    PubMed Central

    Melloni, Chiara; Rao, Sunil V.; Povsic, Thomas J.; Melton, Laura; Kim, Raymond J.; Kilaru, Rakhi; Patel, Manesh; Talan, Mark; Ferrucci, Luigi; Longo, Dan L.; Lakatta, Edward G.; Najjar, Samer S.; Harrington, Robert A.

    2010-01-01

    Background Acute myocardial infarction (MI) remains a leading cause of death despite advances in pharmacologic and percutaneous therapies. Animal models of ischemia/reperfusion have demonstrated that single-dose erythropoietin (EPO) may reduce infarct size, decrease apoptosis, and increase neovascularization, possibly through mobilization of endothelial progenitor cells (EPCs). Study Design REVEAL is a randomized, double-blind, placebo-controlled, multicenter trial evaluating the effects of epoetin alfa on infarct size and left ventricular (LV) remodeling in patients with large MIs. The trial comprises a dose-escalation safety phase and a single-dose efficacy phase using the highest acceptable epoetin alfa dose up to 60,000 units. Up to 250 STEMI patients undergoing primary or rescue percutaneous coronary intervention (PCI) will be randomized to intravenous epoetin alfa or placebo within 4 hours of successful reperfusion. The primary study endpoint is infarct size expressed as a percentage of LV mass, as measured by cardiac magnetic resonance imaging 2–6 days post study medication administration. Secondary endpoints will assess changes in EPC numbers and changes in indices of ventricular remodeling. Conclusion The REVEAL trial will evaluate the safety and efficacy of the highest tolerated single dose of epoetin alfa in patients who have undergone successful rescue or primary PCI for acute STEMI. PMID:21095264

  4. Patients with anaemia can shift from kidney to liver production of erythropoietin as shown by glycoform analysis.

    PubMed

    Lönnberg, Maria; Garle, Mats; Lönnberg, Lina; Birgegård, Gunnar

    2013-01-01

    The primary production site of erythropoietin (EPO) is shifted from the liver to the kidney shortly after birth. Under conditions of lost or reduced kidney production, it is valuable to measure the production capacity of the liver. However, there is a lack of urine or serum based methods that can distinguish endogenous EPO produced in different cell types. Here is presented a method based on chromatographic interaction with the lectin wheat germ agglutinin (WGA) that can distinguish presumably liver-produced EPO, found in anaemic patients receiving epoetin and darbepoetin, from kidney-produced EPO found in healthy individuals. All the tested samples from haemodialysis patients with end-stage renal disease showed a presence of liver EPO. In some samples, the liver-produced EPO made up 90-100% of total EPO at a concentration of 8-10 ng/L in urine, which indicates that the liver has a quite high production capacity, although not adequate for the degree of anaemia. This glycoform analysis has made it possible to affirm that some anaemic patients can increase their liver-production of EPO. The use of such a method can give better insight into the regulation of non-renal endogenous EPO production, a potential source of EPO intended to replace administration of exogenous EPO.

  5. Erythropoietin prevents early and late neuronal demise through modulation of Akt1 and induction of caspase 1, 3, and 8.

    PubMed

    Chong, Zhao Zhong; Lin, Shi-Hua; Kang, Jing-Qiong; Maiese, Kenneth

    2003-03-01

    Erythropoietin (EPO) modulates primarily the proliferation of immature erythroid precursors, but little is known of the potential protective mechanisms of EPO in the central nervous system. We therefore examined the ability of EPO to modulate a series of death-related cellular pathways during anoxia and free radical induced neuronal degeneration. Neuronal injury was evaluated by trypan blue, DNA fragmentation, membrane phosphatidylserine exposure, protein kinase B phosphorylation, cysteine protease activity, mitochondrial membrane potential, and mitogen-activated protein (MAP) kinase phosphorylation. We demonstrate that constitutive neuronal EPO is insufficient to prevent cellular injury, but that signaling through the EPO receptor remains biologically responsive to exogenous EPO administration. Exogenous EPO is both necessary and sufficient to prevent acute genomic DNA destruction and subsequent phagocytosis through membrane PS exposure, because neuronal protection by EPO is completely abolished by co-treatment with an anti-EPO neutralizing antibody. Through pathways that involve the initial activation of protein kinase B, EPO maintains mitochondrial membrane potential. Subsequently, EPO inhibits caspase 8-, caspase 1-, and caspase 3-like activities linked to cytochrome c release through mechanisms that are independent from the MAP kinase systems of p38 and JNK. Elucidating some of the novel neuroprotective pathways employed by EPO may further the development of new therapeutic strategies for neurodegenerative disorders.

  6. Patients with anaemia can shift from kidney to liver production of erythropoietin as shown by glycoform analysis.

    PubMed

    Lönnberg, Maria; Garle, Mats; Lönnberg, Lina; Birgegård, Gunnar

    2013-01-01

    The primary production site of erythropoietin (EPO) is shifted from the liver to the kidney shortly after birth. Under conditions of lost or reduced kidney production, it is valuable to measure the production capacity of the liver. However, there is a lack of urine or serum based methods that can distinguish endogenous EPO produced in different cell types. Here is presented a method based on chromatographic interaction with the lectin wheat germ agglutinin (WGA) that can distinguish presumably liver-produced EPO, found in anaemic patients receiving epoetin and darbepoetin, from kidney-produced EPO found in healthy individuals. All the tested samples from haemodialysis patients with end-stage renal disease showed a presence of liver EPO. In some samples, the liver-produced EPO made up 90-100% of total EPO at a concentration of 8-10 ng/L in urine, which indicates that the liver has a quite high production capacity, although not adequate for the degree of anaemia. This glycoform analysis has made it possible to affirm that some anaemic patients can increase their liver-production of EPO. The use of such a method can give better insight into the regulation of non-renal endogenous EPO production, a potential source of EPO intended to replace administration of exogenous EPO. PMID:23666255

  7. Erythropoietin stimulates spleen BMP4-dependent stress erythropoiesis and partially corrects anemia in a mouse model of generalized inflammation.

    PubMed

    Millot, Sarah; Andrieu, Valérie; Letteron, Philippe; Lyoumi, Saïd; Hurtado-Nedelec, Margarita; Karim, Zoubida; Thibaudeau, Olivier; Bennada, Samira; Charrier, Jean-Luc; Lasocki, Sigismond; Beaumont, Carole

    2010-12-23

    Mouse bone marrow erythropoiesis is homeostatic, whereas after acute anemia, bone morphogenetic protein 4 (BMP4)-dependent stress erythropoiesis develops in the spleen. The aim of this work was to compare spleen stress erythropoiesis and bone marrow erythropoiesis in a mouse model of zymosan-induced generalized inflammation, which induces long-lasting anemia and to evaluate the ability of erythropoietin (Epo) injections to correct anemia in this setting. The effects of zymosan and/or Epo injections on erythroid precursor maturation and apoptosis, serum interferon-γ levels, hematologic parameters, and spleen BMP4 expression were analyzed, as well as the effect of zymosan on red blood cell half-life. We found that bone marrow erythropoiesis is suppressed by inflammation and does not respond to Epo administration, despite repression of erythroblast apoptosis. On the contrary, a robust erythropoietic response takes place in the spleen after Epo injections in both control and zymosan-induced generalized inflammation mice. This specific response implies Epo-mediated induction of BMP4 expression by F4/80(+) spleen macrophages, proliferation of stress burst-forming units-erythroid, and increased number of spleen erythroblasts. It allows only partial recovery of anemia, probably because of peripheral destruction of mature red cells. It is not clear whether similar BMP4-dependent stress erythropoiesis can occur in human bone marrow after Epo injections.

  8. Mathematical modeling reveals differential effects of erythropoietin on proliferation and lineage commitment of human hematopoietic progenitors in early erythroid culture

    PubMed Central

    Ward, Daniel; Carter, Deborah; Homer, Martin; Marucci, Lucia; Gampel, Alexandra

    2016-01-01

    Erythropoietin is essential for the production of mature erythroid cells, promoting both proliferation and survival. Whether erythropoietin and other cytokines can influence lineage commitment of hematopoietic stem and progenitor cells is of significant interest. To study lineage restriction of the common myeloid progenitor to the megakaryocyte/erythroid progenitor of peripheral blood CD34+ cells, we have shown that the cell surface protein CD36 identifies the earliest lineage restricted megakaryocyte/erythroid progenitor. Using this marker and carboxyfluorescein succinimidyl ester to track cell divisions in vitro, we have developed a mathematical model that accurately predicts population dynamics of erythroid culture. Parameters derived from the modeling of cultures without added erythropoietin indicate that the rate of lineage restriction is not affected by erythropoietin. By contrast, megakaryocyte/erythroid progenitor proliferation is sensitive to erythropoietin from the time that CD36 first appears at the cell surface. These results shed new light on the role of erythropoietin in erythropoiesis and provide a powerful tool for further study of hematopoietic progenitor lineage restriction and erythropoiesis. PMID:26589912

  9. Physician Education: The Erythropoietin Receptor and Signal Transduction.

    PubMed

    Yoshimura; Arai

    1996-01-01

    ERYTHROPOIETIN (EPO): Erythropoietin (EPO) is a hormone that promotes the proliferation and differentiation of erythroid progenitor cells and regulates the number of erythrocytes in peripheral blood. EPO is produced mainly by the kidneys, and transcription of the EPO gene is promoted by a reduction in the oxygen concentration in the blood. The existence of EPO was suggested near the end of the 19th century by the discovery that hypoxia increases the production of red blood cells. EPO was identified as a serum factor in the 1950s, and in 1970 Miyake and coworkers succeeded in purifying it by using the urine of patients with aplastic anemia as a starting material. The human EPO gene was cloned in 1985 using a partial amino acid sequence from this purified EPO, and it is well known that recombinant EPO is currently used as a drug to treat anemia associated with chronic renal failure and other illnesses. ACTION OF EPO: When human bone marrow cells are cultured in a semisolid medium containing EPO, they form small erythroblast colonies in five to seven days, and by day 10 large erythroblast colonies appear that resemble fireworks ("burst" colonies). The original cells in the former colonies are called colony forming units-erythroid (CFU-E) or late-stage erythroblast progenitor cells and in the latter colonies they are called burst forming units-erythroid (BFU-E) or early-stage erythroblast progenitor cells. As shown in Figure 1, red blood cells are produced through differentiation from stem cells to BFU-E, CFU-E, and erythroblasts. Although EPO acts on both BFU-E and CFU-E cells, CFU-E cells show greater sensitivity to EPO, and other factors such as stem cell factor (SCF), interleukin (IL)-3, IL-4, and granulocyte macrophage colony-stimulating factor (GM-CSF) must be present together with EPO for BFU-E cell proliferation. In erythroblasts beyond the CFU-E stage, sensitivity to EPO decreases as the cells mature. THE EPO RECEPTOR AND THE CYTOKINE RECEPTOR FAMILY: The EPO

  10. Intravenous Erythropoietin in Patients with ST-Segment Elevation Myocardial Infarction

    PubMed Central

    Najjar, Samer S.; Rao, Sunil V.; Melloni, Chiara; Raman, Subha V.; Povsic, Thomas J.; Melton, Laura; Barsness, Gregory W.; Prather, Kristi; Heitner, John F.; Kilaru, Rakhi; Gruberg, Luis; Hasselblad, Vic; Greenbaum, Adam B.; Patel, Manesh; Kim, Raymond J.; Talan, Mark; Ferrucci, Luigi; Longo, Dan L.; Lakatta, Edward G.; Harrington, Robert A.

    2012-01-01

    Context Acute ST-segment elevation myocardial infarction (STEMI) is a leading cause of morbidity and mortality. In experimental models of MI, erythropoietin reduces infarct size and improves left ventricular (LV) function. Objective To evaluate the safety and efficacy of a single intravenous bolus of epoetin alfa in patients with STEMI. Design, Setting, and Patients Prospective, randomized, double-blind, placebo-controlled trial with a dose-escalation safety phase and a single-dose (60,000 units of epoetin alfa) efficacy phase involving 222 patients with STEMI who underwent successful percutaneous coronary intervention (PCI) as a primary or rescue reperfusion strategy. Intervention Participants were randomly assigned to treatment with intravenous epoetin alfa or matching saline placebo administered within 4 hours of reperfusion. Main Outcome Measure Infarct size, expressed as a percentage of LV mass, assessed by cardiac magnetic resonance (CMR) imaging 2–6 days after study medication administration. Results In the efficacy cohort (n=138), infarct size did not differ between groups at either 2–6 days (15.8±10.3 vs. 15.0±10.0, P=.666) or 12±2 weeks (10.6±8.6 vs. 10.4±7.6, P=.886). Left ventricular ejection fraction also did not differ between groups at either the early (48.2±9.1 vs. 48.9±8.7, P=.671) or late (52.5±9.3 vs. 52.0±8.8, P=.760) timepoints. In pre-specified analyses of patients aged ≥70 years (n=21), mean infarct size within the first week was larger in the epoetin alfa arm than in the placebo group (19.9±9.9 vs.11.7±7.2, P=.026). Patients who received epoetin alfa had a higher incidence of the composite endpoint of death, myocardial infarction, stroke, or stent thrombosis (4.0% vs. 0.0%, P=.042), and a higher incidence of serious adverse events (20.0% vs. 10.3%, P=.052). Conclusions In STEMI patients successfully reperfused with primary or rescue PCI, a single intravenous bolus of epoetin alfa did not reduce infarct size and was

  11. Iron supplementation enhances response to high doses of recombinant human erythropoietin in preterm infants

    PubMed Central

    Carnielli, V.; Da Riol, R.; Montini, G.

    1998-01-01

    AIMS—To determine whether iron supplementation would enhance erythropoiesis in preterm infants treated with high doses of human recombinant erythropoietin (r-HuEPO).
METHODS—Sixty three preterm infants were randomly allocated at birth to one of three groups to receive: r-HuEPO alone, 1200 IU/kg/week (EPO); or r-HuEPO and iron, 1200 IU/kg/week of r-HuEPO plus 20mg/kg/week of intravenous iron (EPO+iron); or to serve as controls. All three groups received blood transfusions according to uniform guidelines.
RESULTS—Infants in the EPO+iron group needed fewer transfusions than controls—mean (95% CI) 1.0 (0.28-1.18) vs 2.9 (1.84-3.88) and received lower volumes of blood—mean (95% CI) 16.7 (4.9-28.6) vs 44.4 (29.0-59.7) ml/kg. The EPO group also needed lower volumes of blood than the controls—mean (95% CI) 20.1 (6.2-34.2) vs 44.4(29.0-59.7) ml/kg, but the same number of transfusions, 1.3 (0.54-2.06) vs 2.9 (1.84-3.88). Reticulocyte and haematocrit values from postnatal weeks 5 to 8 were higher in the EPO+iron than in the EPO group, and both groups had higher values than the controls. Mean (SEM) plasma ferritin was lower in the EPO group—65 (55) µg/l than in the EPO+iron group 780 (182) µg/l, and 561 (228) µg/l in the control infants.
CONCLUSIONS—Early administration of high doses of r-HuEPO with iron supplements significantly reduced the need for blood transfusion. Intravenous iron (20 mg/kg/week in conjunction with r-HuEPO yielded a higher reticulocyte count and haematocrit concentration after the forth week of life than r-HuEPO alone. Infants treated with r-HuEPO alone showed signs of reduced iron stores.

 PMID:9797624

  12. Health-related quality of life in the era of erythropoietin.

    PubMed

    Weisbord, Steven D; Kimmel, Paul L

    2008-01-01

    Patients with end-stage renal disease treated with maintenance hemodialysis suffer substantial impairments in health-related quality of life (HRQOL). Despite widespread efforts, there are few interventions that improve the overall well-being and quality of life of this patient population. The current review provides a description of HRQOL as an essential, yet arguably overlooked health-related domain in hemodialysis patients, and discusses interventions that have been evaluated to improve the functional status and well-being of this population, with a particular focus on therapy with recombinant human erythropoietin. We review the controversy surrounding recombinant human erythropoietin as it relates to HRQOL, and describe the delicate balance faced by renal providers who seek to reduce hemodialysis patients' morbidity and mortality while simultaneously striving to improve patients' HRQOL.

  13. Influence of recombinant human erythropoietin on hematological and hemostatic parameters with special reference to microhemolysis.

    PubMed

    Maurin, N; Fitzner, S; Fritz, H; Gladziwa, U; H agel, J; Stefanidis, I

    1995-03-01

    Twenty chronic hemodialysis patients with renal anemia (hematocrit < 25%) received recombinant human erythropoietin (40 IU/kg body weight 3 x weekly) intravenously after each dialysis. Prior to and at 4, 8 and 12 weeks after commencement of erythropoietin therapy, hematocrit together with hemostasis and microhemolysis parameters were determined. There were significant increases in hematocrit, platelet count and platelet retention, but a significant fall in the initial clearly prolonged bleeding time. Free plasma hemoglobin likewise increased. Conversely, lactate dehydrogenase, prothrombin time, fibrinogen, antithrombin III activity, protein C activity and protein S concentration were all unaltered. The positive effect on bleeding time and platelet retention is most probably caused by an increase in adenosine diphosphate due to the hematocrit-dependent rise in the blood shear stress via physiologic microhemolysis (raised free plasma hemoglobin). PMID:7774078

  14. Erythropoietin protects cardiac myocytes from hypoxia-induced apoptosis through an Akt-dependent pathway.

    PubMed

    Tramontano, Anthony F; Muniyappa, Ranganath; Black, Aislinn D; Blendea, Mihaela C; Cohen, Inna; Deng, Lili; Sowers, James R; Cutaia, Michael V; El-Sherif, Nabil

    2003-09-01

    Apoptosis is a contributing cause of myocyte loss in ischemic heart disease. Recent work has shown that erythropoietin (EPO) offers protection against apoptosis in a wide variety of tissues. We demonstrate that the erythropoietin receptor (EPOR) is expressed in the neonatal rat ventricular myocyte (NRVM). Exposure of NRVMs to hypoxia, with recombinant human EPO, significantly decreased apoptosis as measured by TUNEL, flow cytometry, and caspase 3/7 like activity when compared to hypoxia treatment alone. EPO administered at the initiation of coronary artery occlusion in the rat significantly decreased apoptosis in the myocardial ischemic region. In the NRVM, EPO increased the activity of Akt. The anti-apoptotic effect of EPO was abrogated by co-treatment with LY294002, a specific blocker of phosphatidylinositol 3-kinase (PI3-K). Our study demonstrates that EPO inhibits apoptosis in the NRVM exposed to hypoxia, through an Akt-dependent pathway. EPO also inhibits apoptosis in the in vivo rat model of myocardial ischemia.

  15. "Effects of recombinant human erythropoietin high mimicking abuse doses on oxidative stress processes in rats".

    PubMed

    Bianchi, Sara; Fusi, Jonathan; Franzoni, Ferdinando; Giovannini, Luca; Galetta, Fabio; Mannari, Claudio; Guidotti, Emanuele; Tocchini, Leonardo; Santoro, Gino

    2016-08-01

    Although many studies highlight how long-term moderate dose of Recombinant Human Erythropoietin (rHuEPO) treatments result in beneficial and antioxidants effects, few studies take into account the effects that short-term high doses of rHuEPO (mimicking abuse conditions) might have on the oxidative stress processes. Thus, the aim of this study was to investigate the in vivo antioxidant activity of rHuEPO, administered for a short time and at high doses to mimic its sports abuse as doping. Male Wistar healthy rats (n=36) were recruited for the study and were treated with three different concentrations of rHuEPO: 7.5, 15, 30μg/kg. Plasma concentrations of erythropoietin, 8-epi Prostaglandin F2α, plasma and urinary concentrations of NOx were evaluated with specific assay kit, while hematocrit levels were analyzed with an automated cell counter. Antioxidant activity of rHuEPO was assessed analyzing the possible variation of the plasma scavenger capacity against hydroxylic and peroxylic radicals by TOSC (Total Oxyradical Scavenging Capacity) assay. Statistical analyses showed higher hematocrit values, confirmed by a statistically significant increase of plasmatic EPO concentration. An increase in plasma scavenging capacity against peroxyl and hydroxyl radicals, in 8-isoprostane plasmatic concentrations and in plasmatic and urinary levels of NOX were also found in all the treated animals, though not always statistically significant. Our results confirm the literature data regarding the antioxidant action of erythropoietin administered at low doses and for short times, whereas they showed an opposite incremental oxidative stress action when erythropoietin is administered at high doses. PMID:27470373

  16. [Renal cell carcinoma producing erythrocytosis due to inappropriate production of erythropoietin].

    PubMed

    Villanueva-Gimeno, M M; Vicario-Bermúdez, J M; Fonseca-López, Ch; Caballero-Castro, J P; Zabala-López, S I; Sánchez-Elipe, M A; González-Gómez, N

    2013-01-01

    Erythrocytosis, or polycythaemia, is an increase, in absolute terms, of the erythrocyte mass. The most common solid tumour related to this phenomenon is renal cell carcinoma, which can produce erythrocytosis by increasing erythropoietin production. About 30% of symptomatic renal cell carcinomas are diagnosed due to the appearance of a paraneoplastic syndrome. Polycythaemia is one of these. Surgery, (radical or partial nephrectomy), is the treatment of choice in renal cell carcinoma and helps to keep the erythrocytosis situation under control. PMID:23834980

  17. Changes in erythropoietin levels during space flight or space flight simulation

    NASA Technical Reports Server (NTRS)

    Dunn, C. D. R.; Hen, J. P.

    1980-01-01

    Two hundred and seventy samples from 24 subjects involved in 3 bedrest studies and from 3 subjects involved in Spacelab Mission Development Test 3 were assayed for erythropoietin (Ep), in an in vitro fetal mouse liver cell assay, and for ferritin using a commercially available immunoradiometric assay kit. No trends or significant changes in serum Ep were observed. Serum ferritin concentrations tended to increases slightly during the 'missions', reflecting a redirection of iron from the suppressed erythron into iron stores.

  18. Establishment of the Ph. Eur. erythropoietin chemical reference substance batch 1.

    PubMed

    Burns, C; Bristow, A F; Buchheit, K H; Daas, A; Wierer, M; Costanzo, A

    2015-01-01

    The Erythropoietin (EPO) European Pharmacopoeia (Ph. Eur.) Biological Reference Preparation (BRP) batch 3 was calibrated in 2006 by in vivo bioassay and was used as a reference preparation for these assays as well as for the physicochemical methods in the Ph. Eur. monograph Erythropoietin concentrated solution (1316). In order to avoid the frequent replacement of this standard and thus reduce the use of animals, a new EPO Chemical Reference Substance (CRS) was established to be used solely for the physicochemical methods. Here we report the outcome of a collaborative study aimed at demonstrating the suitability of the candidate CRS (cCRS) as a reference for the physicochemical methods in the Ph. Eur. monograph. Results from the study demonstrated that for the physicochemical methods currently required in the monograph (capillary zone electrophoresis (CZE), polyacrylamide gel electrophoresis (PAGE)/immunoblotting and peptide mapping), the cCRS is essentially identical to the existing BRP. However, data also indicated that, for the physicochemical methods under consideration for inclusion in a revised monograph (test for oxidised forms and glycan mapping), the suitability of the cCRS as a reference needs to be confirmed with additional work. Further to completion of the study, the Ph. Eur. Commission adopted the cCRS as "Erythropoietin for physicochemical tests CRS batch 1" to be used for CZE, PAGE/immunoblotting and peptide mapping.

  19. Reactivity of patients with maintenance hemodialysis to erythropoietin in the treatment of renal anemia.

    PubMed

    Hu, J P; Cheng, X; Xu, X F; Yu, G J; Luo, F; Zhang, G S; Yang, N; Shen, P; Yan, X Y

    2016-01-01

    To explore the reactivity of patients with renal anemia (MHD) to erythropoietin (EPO) in maintenance hemodialysis (HD), 31 patients were enrolled in this study. According to the level of serum ferritin (SF), they were divided into two groups; one group received treatment using recombinant human erythropoietin (rHuEPO) and the other group was given iron sucrose. Taking terminal EPO dosage, terminal erythropoietin resistance index (ERI) and rate of change of ERI (ΔERII) as target indexes, the influence of SF level on dosage of EPO was evaluated after usage conditions of relevant substances in a 3-month period. The results revealed that differences of dialysis age, albumin (ALB), blood calcium, initial and terminal SF, variable quantity of hemoglobin (Hb), terminal EPO and ERI between two groups had statistical significance. Furthermore, SF level and terminal EPO (r = -0.37, P < 0. 05) as well as SF level and terminal ERI (r = - 0.39, P <0.05) were negatively correlated. Difference of terminal ERI between the two groups had statistical significance. It can therefore be summarized that supplementing an iron agent intravenously to maintain SF level between 500 ng/ml and 1200 ng/ml may improve reactivity of patients with MHD to EPO. In addition, rHuEPO therapy in treating anemia of patients with MHD has the same effect with intravenous drug delivery, less side effects and is easy to administer. PMID:27358145

  20. Effects of vitamin E supplementation during erythropoietin treatment of the anaemia of prematurity

    PubMed Central

    Pathak, A; Roth, P; Piscitelli, J; Johnson, L

    2003-01-01

    Aims: To evaluate the effects of vitamin E supplementation on haemoglobin concentration and the requirement for transfusion in premature infants treated with erythropoietin and iron. Methods: Randomised, double blind, placebo controlled trial. Thirty infants ≤32 weeks gestation and ≤1250 g birth weight, who were defined as stable based on minimal requirements for respiratory support and phlebotomy, and absence of major congenital anomalies were enrolled. All were treated with erythropoietin and iron, and were randomised to receive, in addition, either vitamin E 50 IU/day or placebo for eight weeks or until discharge, whichever came first. Results: Despite higher vitamin E (α-tocopherol) levels in the experimental group in weeks 3 (49.0 v 28.1 µmol/l) and 8 (66.2 v 38.5 µmol/l), there were no differences in haemoglobin, reticulocyte count, iron concentration, or transfusion requirement. Conclusions: Oral vitamin E supplementation at 50 IU/day does not increase the response of preterm infants to erythropoietin and iron. Vitamin E obtained through standard nutrition may have been sufficient or higher doses may be required. PMID:12819167

  1. New insights into erythropoietin and epoetin alfa: mechanisms of action, target tissues, and clinical applications.

    PubMed

    Weiss, Mitchell J

    2003-01-01

    Recombinant human erythropoietin (epoetin alfa) has proven beneficial for the treatment of various anemias. The mechanism of action of endogenous erythropoietin and the therapeutic use of epoetin alfa to stimulate red blood cell production and improve the quality of life in cancer patients are reviewed here. Epoetin alfa may also attenuate the cognitive dysfunction associated with cancer therapy. Interestingly, functional endogenous erythropoietin receptor signaling pathways have been demonstrated in numerous nonerythropoietic tissues. Of particular importance, epoetin alfa confers neurotrophic and neuroprotective effects in cultured neurons and in several animal models for neurologic disease. In one clinical trial, epoetin alfa appeared to limit functional and histologic damage in patients with stroke. Therefore, in cancer patients receiving chemotherapy, the beneficial effects of epoetin alfa could be mediated not only through enhanced erythrocyte production but also via direct effects on the nervous system. Further investigation into the nonerythropoietic effects of epoetin alfa could broaden its clinical utility for patients with cancer and also provide new therapies for various neurologic disorders.

  2. Expression of GPI anchored human recombinant erythropoietin in CHO cells is devoid of glycosylation heterogeneity.

    PubMed

    Singh, Pankaj Kumar; Devasahayam, Mercy; Devi, Sobita

    2015-04-01

    Erythropoietin is a glycohormone involved in the regulation of the blood cell levels. It is a 166 amino acid protein having 3 N-glycosylation and one O-linked glycosylation sites, and is used to treat anaemia related illness. Though human recombinant erythropoietin (rEPO) is produced in CHO cells, the loss in quality control is 80% due to incomplete glycosylation of the rEPO with low levels of fully glycosylated active rEPO. Here, we describe the expression from CHO cells of fully glycosylated human rEPO when expressed as a GPI anchored molecule (rEPO-g). The results demonstrated the production of a homogenous completely glycosylated human rEPO-g as a 42 kD band without any low molecular weight glycoform variants as shown by affinity chromatography followed by SDS-PAGE and anti-human EPO specific western blot. The western blot using specific monoclonal antibody is the available biochemical technique to prove the presence of homogeneity in the expressed recombinant protein. The GPI anchor can be removed during the purification process to yield a therapeutically relevant recombinant erythropoietin molecule cells with a higher in vivo biological activity due to its high molecular weight of 40 kD. This is possibly the first report on the production of a homogenous and completely glycosylated human rEPO from CHO cells for efficient therapy. PMID:26011979

  3. Combination Erythropoietin-Hydroxyurea Therapy in Sickle Cell Disease: NIH experience and literature review

    PubMed Central

    Little, Jane A.; McGowan, Vicki R.; Kato, Gregory J.; Partovi, Kristine S.; Feld, Jordan J.; Maric, Irina; Martyr, Sabrina; Taylor, James G.; Machado, Roberto F.; Heller, Theo; Castro, Oswaldo; Gladwin, Mark T.

    2012-01-01

    Erythropoietin is being used more widely in the management of sickle cell disease (SCD, inclusive of homozygous sickle beta, SS, and compound heterozygous sickle beta thalassemia, Sβ0 thal), often in conjunction with hydroxyurea (HU). Herein, we summarize the published experience with erythropoietin use in SCD, including 39 patients (SS, n=30; Sβ0 thal, n=9) who were treated between 1990 and 1996; to which we add 13 patients with Sickle Syndromes (SS, n=12, compound heterozygous SC disease, n=1) who were treated with erythropoietin or darbopoietin (here cumulatively referred to as EPO) at the National Institutes of Health (NIH) since 2002. The dose range of erythropoietin for SCD in the published series, at a median of >200 Units/Kg/dose, is higher than is used in end-stage renal disease. The median duration of erythropoietin therapy in the published series was ≥3 months, with minimal reported side-effects. At the NIH, the median age of Sickle Syndrome patients who received EPO was 51 (24 to 70) years; 12/13 patient had sickle-associated pulmonary hypertension. 11/13 patients were treated with both HU and EPO for > 4 months (median of 11 months on EPO) without complication. 5/13 patients (all HbSS) with pulmonary hypertension were given EPO for reticulocytopenia (<100,000/μL) on HU; 5/13 patients (all HbSS), with pulmonary hypertension, were given EPO and HU concurrently, in light of an estimated glomerular filtration rate of <80 ml/minute. 3/13 patients (2 Hb SS, 1 HbSC) were treated with EPO for miscellaneous reasons. Hematologic responses, detailed herein, were promising. Our experience suggests that EPO may be safe in SCD, when used in conjunction with HU; EPO therapy could allow more aggressive HU dosing in high-risk SCD patients and in the setting of mild renal insufficiency, common to the aging sickle cell population. We outline our current therapeutic strategy for EPO use in SCD. PMID:16885048

  4. [Sequential treatment with granulocyte-colony stimulating factor (GCSF) and human recombinant erythropoietin (rH-EPO) in anemia of a patient with myelodysplastic syndrome and high blood transfusion requirements)].

    PubMed

    Borbolla-Escoboza, J R; González-Avante, C M; López-Hernández, M A; Flores-Chapa, J D; Collados-Larumbe, M T

    1997-01-01

    We describe the case of a patient with myelodysplastic syndrome (MDS) classified as Refractory Anemia with our Excess blasts, who suffered from high transfusional requirements and who did not respond to the administrations of B12 vitamin, folates, danazol, low dose cytarabine or recombinant human erythropoietin (rHuEPO). The patient was administered two cytokines: granulocyte colony stimulating factor (G-CSF) followed by rHuEPO. The patient remained transfusion free for more than 4 months until his death from causes not related to MDS or the therapy he received. It is the opinion of the authors that the initial G-CSF administration stimulated the early erythroid precursors, making them capable of finishing their maturation when rHuEPO was administered. We believe that this could be a useful therapeutic measure in the treatment of patients with MDS and high transfusional requirements.

  5. Administrative Synergy

    ERIC Educational Resources Information Center

    Hewitt, Kimberly Kappler; Weckstein, Daniel K.

    2012-01-01

    One of the biggest obstacles to overcome in creating and sustaining an administrative professional learning community (PLC) is time. Administrators are constantly deluged by the tyranny of the urgent. It is a Herculean task to carve out time for PLCs, but it is imperative to do so. In this article, the authors describe how an administrative PLC…

  6. Preclinical and Preliminary Clinical Evaluation of Genetically Transduced Dermal Tissue Implants for the Sustained Secretion of Erythropoietin and Interferon α.

    PubMed

    Shapir, Nir; Miari, Reem; Blum, Shany; Schwartz, Doron; Chernin, Gil; Neil, Garry A; Afik, Daniel; Panet, Amos

    2015-12-01

    Protein drugs are currently delivered by bolus injection and although treatment frequently is successful, these methods also have major drawbacks, which call for the development of alternative technologies allowing prolonged delivery of these drugs. We developed a new ex vivo gene therapy platform called Transduced Autologous Restorative Gene Therapy (TARGT) for sustained long term production and secretion of autologous therapeutic proteins. A biopsy of dermal tissue taken from the patient is transduced ex vivo with a viral vector encoding the required gene under a constitutive promoter. Following measurement of protein secretion ex vivo, the transduced dermal tissue is implanted back into the patient, where it secretes the therapeutic protein into the circulation for several months or longer. A major hurdle to this approach is potential immunogenicity of the transduced tissue following implantation. In this paper we describe the preclinical and early clinical development of this technology, which allowed for overcoming these hurdles. To that end, we have used the helper dependent (HD) adenoviral vector with newly designed expression cassette containing genetic elements to optimize transgene expression. Moreover, we have developed procedures for TARGT tissue implantation, with measures to improve engraftment and reduce inflammation and rejection. Implantation of human TARGT to severe combined immune deficient (SCID) mice indicated long-term production of active proteins in the blood. Preliminary results of a clinical trial from two anemic end-stage renal disease patients, implanted with TARGTs expressing the human erythropoietin (EPO) gene, demonstrated prolonged secretion with physiologic blood level of the hormone and hemoglobin maintenance in the desired range, for a period of at least 5 months without exogenous EPO administration. We believe that the TARGT technology has the potential to become a platform for the sustained delivery of therapeutic proteins in

  7. A single topical dose of erythropoietin applied on a collagen carrier enhances calvarial bone healing in pigs

    PubMed Central

    2014-01-01

    Background and purpose The osteogenic potency of erythropoietin (EPO) has been documented. However, its efficacy in a large-animal model has not yet been investigated; nor has a clinically safe dosage. The purpose of this study was to overcome such limitations of previous studies and thereby pave the way for possible clinical application. Our hypothesis was that EPO increases calvarial bone healing compared to a saline control in the same subject. Methods We used a porcine calvarial defect model. In each of 18 pigs, 6 cylindrical defects (diameter: 1 cm; height: 1 cm) were drilled, allowing 3 pairwise comparisons. Treatment consisted of either 900 IU/mL EPO or an equal volume of saline in combination with either autograft, a collagen carrier, or a polycaprolactone (PCL) scaffold. After an observation time of 5 weeks, the primary outcome (bone volume fraction (BV/TV)) was assessed with high-resolution quantitative computed tomography. Secondary outcome measures were histomorphometry and blood samples. Results The median BV/TV ratio of the EPO-treated collagen group was 1.06 (CI: 1.02–1.11) relative to the saline-treated collagen group. Histomorphometry showed a similar median effect size, but it did not reach statistical significance. Autograft treatment had excellent healing potential and was able to completely regenerate the bone defect independently of EPO treatment. Bony ingrowth into the PCL scaffold was sparse, both with and without EPO. Neither a substantial systemic effect nor adverse events were observed. The number of blood vessels was similar in EPO-treated defects and saline-treated defects. Interpretation Topical administration of EPO on a collagen carrier moderately increased bone healing. The dosing regime was safe, and could have possible application in the clinical setting. However, in order to increase the clinical relevance, a more potent but still clinically safe dose should be investigated. PMID:24564750

  8. Mimicking hypoxia to treat anemia: HIF-stabilizer BAY 85-3934 (Molidustat) stimulates erythropoietin production without hypertensive effects.

    PubMed

    Flamme, Ingo; Oehme, Felix; Ellinghaus, Peter; Jeske, Mario; Keldenich, Jörg; Thuss, Uwe

    2014-01-01

    Oxygen sensing by hypoxia-inducible factor prolyl hydroxylases (HIF-PHs) is the dominant regulatory mechanism of erythropoietin (EPO) expression. In chronic kidney disease (CKD), impaired EPO expression causes anemia, which can be treated by supplementation with recombinant human EPO (rhEPO). However, treatment can result in rhEPO levels greatly exceeding the normal physiological range for endogenous EPO, and there is evidence that this contributes to hypertension in patients with CKD. Mimicking hypoxia by inhibiting HIF-PHs, thereby stabilizing HIF, is a novel treatment concept for restoring endogenous EPO production. HIF stabilization by oral administration of the HIF-PH inhibitor BAY 85-3934 (molidustat) resulted in dose-dependent production of EPO in healthy Wistar rats and cynomolgus monkeys. In repeat oral dosing of BAY 85-3934, hemoglobin levels were increased compared with animals that received vehicle, while endogenous EPO remained within the normal physiological range. BAY 85-3934 therapy was also effective in the treatment of renal anemia in rats with impaired kidney function and, unlike treatment with rhEPO, resulted in normalization of hypertensive blood pressure in a rat model of CKD. Notably, unlike treatment with the antihypertensive enalapril, the blood pressure normalization was achieved without a compensatory activation of the renin-angiotensin system. Thus, BAY 85-3934 may provide an approach to the treatment of anemia in patients with CKD, without the increased risk of adverse cardiovascular effects seen for patients treated with rhEPO. Clinical studies are ongoing to investigate the effects of BAY 85-3934 therapy in patients with renal anemia. PMID:25392999

  9. Activation of aryl hydrocarbon receptor mediates suppression of hypoxia-inducible factor-dependent erythropoietin expression by indoxyl sulfate.

    PubMed

    Asai, Hirobumi; Hirata, Junya; Hirano, Ayumi; Hirai, Kazuya; Seki, Sayaka; Watanabe-Akanuma, Mie

    2016-01-15

    Indoxyl sulfate (IS) is a representative uremic toxin that accumulates in the blood of patients with chronic kidney disease (CKD). In addition to the involvement in the progression of CKD, a recent report indicates that IS suppresses hypoxia-inducible factor (HIF)-dependent erythropoietin (EPO) production, suggesting that IS may also contribute to the progression of renal anemia. In this report, we provide evidence that aryl hydrocarbon receptor (AhR) mediates IS-induced suppression of HIF activation and subsequent EPO production. In HepG2 cells, IS at concentrations similar to the blood levels in CKD patients suppressed hypoxia- or cobalt chloride-induced EPO mRNA expression and transcriptional activation of HIF. IS also induced AhR activation, and AhR blockade resulted in abolishment of IS-induced suppression of HIF activation. The HIF transcription factor is a heterodimeric complex composed of HIF-α subunits (HIF-1α and HIF-2α) and AhR nuclear translocator (ARNT). IS suppressed nuclear accumulation of the HIF-α-ARNT complex accompanied by an increase of the AhR-ARNT complex in the nucleus, implying the involvement of interactions among AhR, HIF-α, and ARNT in the suppression mechanism. In rats, oral administration of indole, a metabolic precursor of IS, inhibited bleeding-induced elevation of renal EPO mRNA expression and plasma EPO concentration and strongly induced AhR activation in the liver and renal cortex tissues. Collectively, this study is the first to elucidate the detailed mechanism by which AhR plays an indispensable role in the suppression of HIF activation by IS. Hence, IS-induced activation of AhR may be a potential therapeutic target for treating renal anemia. PMID:26561638

  10. Early adoption of cyclosporine and recombinant human erythropoietin: clinical, economic, and policy issues with emergence of high-cost drugs.

    PubMed

    Powe, N R; Eggers, P W; Johnson, C B

    1994-07-01

    The discovery of new drugs and their introduction into US markets will become an intense area of focus should health care reform result in Medicare insurance coverage for prescription drugs. Particular attention will be focused on high-cost drugs. Two high-cost drugs, cyclosporine and recombinant human erythropoietin (rHuEPO), introduced into the clinical management of patients with kidney disease during the past decade, provide some experience concerning the forces affecting the use of expensive drugs in a cost-conscious health care system. The decision to prescribe a drug will depend on provider's judgements of the drug's clinical benefits and costs compared with those of other possible therapies. It may also depend on payment policy. Both cyclosporine and rHuEPO were adopted rapidly and extensively by providers of end-stage renal disease care following US Food and Drug Administration approval, despite their high costs. Both drugs were remarkably effective, relatively safe, and able to be administered without great difficulty compared with the therapies they have replaced. There was no additional payment to hospitals for the initial use of cyclosporine, which was introduced in 1983 at the time when Medicare's prospective payment was established, since choice of immunosuppressive agent did not affect the fixed, per-admission payment determined by the diagnosis-related group for kidney transplantation. Medicare coverage for continuing outpatient use of cyclosporine was not initially provided, in contrast to rHuEPO, which was introduced in 1989 with Medicare outpatient coverage and payment of 80% of the allowed charge. Despite their high costs and different methods of insurance payment both drugs achieved a rather quick and high penetration rate into their respective populations.(ABSTRACT TRUNCATED AT 250 WORDS)

  11. A Review of Safety, Efficacy, and Utilization of Erythropoietin, Darbepoetin, and Peginesatide for Patients with Cancer or Chronic Kidney Disease: A Report from the Southern Network on Adverse Reactions (SONAR)

    PubMed Central

    Bennett, Charles L.; Spiegel, David M.; Macdougall, Iain C.; Norris, LeAnn; Qureshi, Zaina P.; Sartor, Oliver; Lai, Stephen Y.; Tallman, Martin S.; Raisch, Dennis W.; Smith, Sheila Weiss; Silver, Samuel; Murday, Alanna S.; Armitage, James O.; Goldsmith, David

    2014-01-01

    The erythropoiesis-stimulating agents (ESAs) erythropoietin and darbepoetin prevent transfusions among chemotherapy-associated anemia patients. Clinical trials, meta-analyses, and guidelines identify mortality, tumor progression, and venous thromboembolism (VTE) risks with ESA administration in this setting. Product labels advise against administering ESAs with potentially curative chemotherapy (United States) or to conduct risk–benefit assessments (Europe/Canada). Since 2007, fewer chemotherapy-associated anemia patients in the United States and Europe receive ESAs. ESAs and the erythropoietin receptor agonist peginesatide prevent transfusions among chronic kidney disease (CKD) patients; clinical trials, guidelines, and meta-analyses demonstrate myocardial infarction, stroke, VTE, or mortality risks with ESAs targeting high hemoglobin levels. U.S. labels recommend administering ESAs or peginesatide at doses sufficient to prevent transfusions among dialysis CKD patients. For dialysis CKD patients, Canadian and European labels recommend targeting hemoglobin levels of 10 to 12 g/dL and 11 to 12 g/dL, respectively, with ESAs. ESA utilization for dialysis CKD patients has decreased in the United States. PMID:23111861

  12. A review of safety, efficacy, and utilization of erythropoietin, darbepoetin, and peginesatide for patients with cancer or chronic kidney disease: a report from the Southern Network on Adverse Reactions (SONAR).

    PubMed

    Bennett, Charles L; Spiegel, David M; Macdougall, Iain C; Norris, LeAnn; Qureshi, Zaina P; Sartor, Oliver; Lai, Stephen Y; Tallman, Martin S; Raisch, Dennis W; Smith, Sheila Weiss; Silver, Samuel; Murday, Alanna S; Armitage, James O; Goldsmith, David

    2012-11-01

    The erythropoiesis-stimulating agents (ESAs) erythropoietin and darbepoetin prevent transfusions among chemotherapy-associated anemia patients. Clinical trials, meta-analyses, and guidelines identify mortality, tumor progression, and venous thromboembolism (VTE) risks with ESA administration in this setting. Product labels advise against administering ESAs with potentially curative chemotherapy (United States) or to conduct risk-benefit assessments (Europe/Canada). Since 2007, fewer chemotherapy-associated anemia patients in the United States and Europe receive ESAs. ESAs and the erythropoietin receptor agonist peginesatide prevent transfusions among chronic kidney disease (CKD) patients; clinical trials, guidelines, and meta-analyses demonstrate myocardial infarction, stroke, VTE, or mortality risks with ESAs targeting high hemoglobin levels. U.S. labels recommend administering ESAs or peginesatide at doses sufficient to prevent transfusions among dialysis CKD patients. For dialysis CKD patients, Canadian and European labels recommend targeting hemoglobin levels of 10 to 12 g/dL and 11 to 12 g/dL, respectively, with ESAs. ESA utilization for dialysis CKD patients has decreased in the United States.

  13. Activating mitochondrial function and haemoglobin expression with EH-201, an inducer of erythropoietin in neuronal cells, reverses memory impairment

    PubMed Central

    Horng, Lin-Yea; Hsu, Pei-Lun; Chen, Li-Wen; Tseng, Wang-Zou; Hsu, Kai-Tin; Wu, Chia-Ling; Wu, Rong-Tsun

    2015-01-01

    Background and Purpose Memory impairment can be progressive in neurodegenerative diseases, and physiological ageing or brain injury, mitochondrial dysfunction and oxidative stress are critical components of these issues. An early clinical study has demonstrated cognitive improvement during erythropoietin treatment in patients with chronic renal failure. As erythropoietin cannot freely cross the blood–brain barrier, we tested EH-201 (2,3,5,4′-tetrahydroxystilbene-2-O-β-d-glucoside, also known as TSG), a low MW inducer of erythropoietin, for its therapeutic effects on memory impairment in models of neurodegenerative diseases, physiological ageing or brain injury. Experimental Approach The effects of EH-201 were investigated in astrocytes and PC12 neuronal-like cells. In vivo, we used sleep-deprived (SD) mice as a stress model, amyloid-β (Aβ)-injected mice as a physiological ageing model and kainic acid (KA)-injected mice as a brain damage model to assess the therapeutic effects of EH-201. Key Results EH-201 induced expression of erythropoietin, PPAR-γ coactivator 1α (PGC-1α) and haemoglobin in astrocytes and PC12 neuronal-like cells. In vivo, EH-201 treatment restored memory impairment, as assessed by the passive avoidance test, in SD, Aβ and KA mouse models. In the hippocampus of mice given EH-201 in their diet, levels of erythropoietin, PGC-1α and haemoglobin were increased Conclusions and Implications The induction of endogenous erythropoietin in neuronal cells by inducers such as EH-201 might be a therapeutic strategy for memory impairment in neurodegenerative disease, physiological ageing or traumatic brain injury. PMID:26177968

  14. Is the haematopoietic effect of testosterone mediated by erythropoietin? The results of a clinical trial in older men.

    PubMed

    Maggio, M; Snyder, P J; Ceda, G P; Milaneschi, Y; Luci, M; Cattabiani, C; Masoni, S; Vignali, A; Volpi, R; Lauretani, F; Peachey, H; Valenti, G; Cappola, A R; Longo, D; Ferrucci, L

    2013-01-01

    The stimulatory effects of testosterone on erythropoiesis are very well known, but the mechanisms underlying the erythropoietic action of testosterone are still poorly understood, although erythropoietin has long been considered a potential mediator. A total of 108 healthy men >65 years old with serum testosterone concentration <475 ng/dL were recruited by direct mailings to alumni of the University of Pennsylvania and Temple University, and randomized to receive a 60-cm(2) testosterone or placebo patch for 36 months. Ninety-six subjects completed the trial. We used information and stored serum specimens from this trial to test the hypothesis that increasing testosterone increases haemoglobin by stimulating erythropoietin production. We used information of 67 men, 43 in the testosterone group and 24 in the placebo group who had banked specimens available for assays of testosterone, haemoglobin and erythropoietin at baseline and after 36 months. The original randomized clinical study was primarily designed to verify the effects of testosterone on bone mineral density. The primary outcome of this report was to investigate whether or not transdermal testosterone increases haemoglobin by increasing erythropoietin levels. The mean age ± SD of the 67 subjects at baseline was 71.8 ± 4.9 years. Testosterone replacement therapy for 36 months, as compared with placebo, induced a significant increase in haemoglobin (0.86 ± 0.31 g/dL, p = 0.01), but no change in erythropoietin levels (-0.24 ± 2.16 mIU/mL, p = 0.91). Included time-varying measure of erythropoietin did not significantly account for the effect of testosterone on haemoglobin (Treatment-by-time: β = 0.93, SE = 0.33, p = 0.01). No serious adverse effect was observed. Transdermal testosterone treatment of older men for 36 months significantly increased haemoglobin, but not erythropoietin levels. The haematopoietic effect of testosterone does not appear to be mediated by stimulation of

  15. Oxidative stress during erythropoietin hyporesponsiveness anemia at end stage renal disease: Molecular and biochemical studies

    PubMed Central

    Khalil, Samar K.M.; Amer, H.A.; El Behairy, Adel M.; Warda, Mohamad

    2016-01-01

    Inflammation and oxidative stress are two faces of one coin in end stage renal disease patients (ESRD) on maintenance hemodialysis. Their interconnection induces anemia complicated with erythropoietin hyporesponsiveness. The biochemical bases behind the resistance to erythropoietin therapy with frequent hemoglobinemia, oxidative stress and iron status have not been fully understood. Here two equal groups (40 patients each) of responders and non-responders to recombinant human erythropoietin therapy (higher than 300 IU/kg/wk of epoetin) were investigated. Hematological and biochemical analyses of collected blood and serum samples were performed along with serum electrophoretic protein footprinting. The leukocytic DNA fragmentation was used to evaluate the degree of oxidative insult. The good responders showed lower erythrocyte malondialdehyde (E-MDA) level and less DNA fragmentation of circulating leukocytes than poor responders with elevated hemoglobin, albumin, A/G ratio, total iron, and ferritin levels. Contrariwise, lower erythrocyte superoxide dismutase (E-SOD) and catalase activities in EPO poor responder group were noticed. Neither other serum constituents nor electrophoretic protein pattern showed any difference between the two groups. There were higher levels of inflammatory markers, interleukin-6 (IL6) and C-reactive protein (CRP) in EPO poor responder than good responder. The negative correlations between Hb and both IL6 and CRP levels in the present data remotely indicate a positive correlation between inflammatory markers and severity of anemia. A direct correlation between Hb and antioxidant enzymes (E-SOD and catalase) was noticed, while inverse correlation with E-MDA was recorded. The study proved that oral supplementation of vitamin C to ESRD patients might mitigate the previously elevated serum MDA level in these patients. PMID:27222740

  16. Erythropoietin improves synaptic transmission during and following ischemia in rat hippocampal slice cultures.

    PubMed

    Weber, Astrid; Maier, Rolf F; Hoffmann, Ulrike; Grips, Martin; Hoppenz, Marc; Aktas, Ayse G; Heinemann, Uwe; Obladen, Michael; Schuchmann, Sebastian

    2002-12-27

    Erythropoietin (EPO) prevents neuronal damage following ischemic, metabolic, and excitotoxic stress. In this study evoked extracellular field potentials (FP) were used to investigate the effect of EPO on synaptic transmission in hippocampal slice cultures. EPO treated cultured slices (40 units/ml for 48 h) showed significantly increased FP during and following oxygen and glucose deprivation compared with untreated control slices. The addition of the Jak2 inhibitor AG490 (50 microM for 48 h) blocked the EPO effect. These data suggest that EPO improves synaptic transmission during and following ischemia in hippocampal slice cultures.

  17. [Erythropoietin-forming and esterase activity of rat kidney subcellular fractions during stimulation of erythropoiesis].

    PubMed

    Novikov, N M; Voronkov, S F; Voloshchenko, L G; Mikhaĭlova, S N

    1977-01-01

    Stimulation of erythropoiesis in rats (hemolytic-phenylhydrazine and acute posthemorrhagic anemia, effect of hypoxic hypoxia) was accompanied by an increased erythropoietine-formating activity in kidney microsomes and light mitochondria. The phenomenon correlated with an increased esterase activity in hypotonic supernatant of kidney homogenate mainly due to the enzymatic fraction, corresponding to alpha2-globulin by its mobility. Histochemical examination of kidney showed that the most distinct alterations in the esterase activity were observed in epithelial cells of nephron proximal part and in capillary endothelium.

  18. Administrative Ecology

    ERIC Educational Resources Information Center

    McGarity, Augustus C., III; Maulding, Wanda

    2007-01-01

    This article discusses how all four facets of administrative ecology help dispel the claims about the "impossibility" of the superintendency. These are personal ecology, professional ecology, organizational ecology, and community ecology. Using today's superintendency as an administrative platform, current literature describes a preponderance of…

  19. Administrative Support.

    ERIC Educational Resources Information Center

    Doran, Dorothy; And Others

    This guide is intended to assist business education teachers in administrative support courses. The materials presented are based on the Arizona validated occupational competencies and tasks for the occupations of receptionist, secretary, and administrative assistant. Word processing skills have been infused into each of the three sections. The…

  20. White matter changes in patients with Friedreich ataxia after treatment with erythropoietin

    PubMed Central

    Egger, Karl; Clemm von Hohenberg, Christian; Schocke, Michael F; Guttmann, Charles RG; Wassermann, Demian; Wigand, Marlene C; Nachbauer, Wolfgang; Kremser, Christian; Sturm, Brigitte; Scheiber-Mojdehkar, Barbara; Kubicki, Marek; Shenton, Martha E; Boesch, Sylvia

    2013-01-01

    Background and Purpose Erythropoietin (EPO) has received growing attention because of its neuro-regenerative properties. Preclinical and clinical evidence supports its therapeutic potential in brain conditions like stroke, multiple sclerosis and schizophrenia. Also in Friedreich ataxia, clinical improvement after EPO therapy was shown. The aim of the present study was to assess possible therapy-associated brain white-matter changes in these patients. Methods Nine patients with Friedreich ataxia underwent Diffusion Tensor Imaging (DTI) before and after EPO treatment. Tract-based spatial statistics (TBSS) was used for longitudinal comparison. Results We detected widespread longitudinal increase in fractional anisotropy (FA) and axial diffusivity (D||) in cerebral hemispheres bilaterally (p<0.05, corrected), while no changes were observed within the cerebellum, medulla oblongata and pons. Conclusions To the best of our knowledge, this is the first DTI study to investigate the effects of erythropoietin in a neurodegenerative disease. Anatomically, the diffusivity changes appear disease-unspecific, and their biological underpinnings deserve further study. PMID:24015771

  1. Exploring Erythropoietin and G-CSF Combination Therapy in Chronic Stroke Patients

    PubMed Central

    Shin, Yoon-Kyum; Cho, Sung-Rae

    2016-01-01

    Erythropoietin (EPO) and granulocyte-colony stimulating factor (G-CSF) are known to have neuroprotective actions. Based on previous reports showing the synergistic effects of EPO+G-CSF combination therapy in experimental models, we investigated the safety of EPO+G-CSF combination therapy in patients with chronic stroke. In a pilot study, 3 patients were treated with EPO and G-CSF for 5 consecutive days, with follow-up on day 30. In an exploratory double-blind study, 6 patients were allocated to treatment with either EPO+G-CSF or placebo. Treatment was applied once a day for 5 days per month over 3 months. Participants were followed up for 6 months. To substantiate safety, vital signs, adverse events, and hematological values were measured on days 0, 5, and 30 in each cycle and on day 180. Functional outcomes were determined on day 0 and 180. In the laboratory measurements, EPO+G-CSF combination therapy significantly elevated erythropoietin, CD34+ hematopoietic stem cells, white blood cells, and neutrophils on day 5 of each cycle. There were no observations of serious adverse events. In the functional outcomes, the grip power of the dominant hand was increased in the EPO+G-CSF treatment group. In conclusion, this exploratory study suggests a novel strategy of EPO+G-CSF combination therapy for stroke patients. PMID:27043535

  2. Exploring Erythropoietin and G-CSF Combination Therapy in Chronic Stroke Patients.

    PubMed

    Shin, Yoon-Kyum; Cho, Sung-Rae

    2016-01-01

    Erythropoietin (EPO) and granulocyte-colony stimulating factor (G-CSF) are known to have neuroprotective actions. Based on previous reports showing the synergistic effects of EPO+G-CSF combination therapy in experimental models, we investigated the safety of EPO+G-CSF combination therapy in patients with chronic stroke. In a pilot study, 3 patients were treated with EPO and G-CSF for 5 consecutive days, with follow-up on day 30. In an exploratory double-blind study, 6 patients were allocated to treatment with either EPO+G-CSF or placebo. Treatment was applied once a day for 5 days per month over 3 months. Participants were followed up for 6 months. To substantiate safety, vital signs, adverse events, and hematological values were measured on days 0, 5, and 30 in each cycle and on day 180. Functional outcomes were determined on day 0 and 180. In the laboratory measurements, EPO+G-CSF combination therapy significantly elevated erythropoietin, CD34⁺ hematopoietic stem cells, white blood cells, and neutrophils on day 5 of each cycle. There were no observations of serious adverse events. In the functional outcomes, the grip power of the dominant hand was increased in the EPO+G-CSF treatment group. In conclusion, this exploratory study suggests a novel strategy of EPO+G-CSF combination therapy for stroke patients. PMID:27043535

  3. Factors related to transfusion in very low birthweight infants treated with erythropoietin.

    PubMed Central

    Maier, R. F.; Obladen, M.; Messinger, D.; Wardrop, C. A.

    1996-01-01

    The need for red cell transfusions is reduced but not eliminated by recombinant human erythropoietin (rhEPO) in very low birthweight (VLBW) infants. To detect factors associated with the decision to transfuse VLBW infants during rhEPO treatment and to explain rhEPO 'non-responders', the subgroup of those 120 VLBW infants who were treated with rhEPO 750 IU/kg per week in the second European Multicentre rhEPO Trial was evaluated. Sixty (50%) infants received at least one transfusion during erythropoietin treatment. Transfusion was frequent in infants with extremely low birthweight (79% for 750-999 g), low gestational age (70% for < or = 28 weeks), low initial haematocrit or low initial reticulocyte count (61% for haematocrit < or = 0.48 and reticulocytes < or = 9%, respectively). Considerable differences among centres were found for sampling blood loss, iron supply, and transfusion rate, which ranged from 13% to 73% and was related to the volume of diagnostic blood loss (19% vs 80% for blood loss < 1 vs > or = 1 ml/kg per day). The prognostic variables birthweight, initial haematocrit, and gestational age were found to be most predictive for transfusion. To improve rhEPO response in VLBW infants, there is a need to minimise diagnostic blood loss, to prevent iron deficiency, and to develop rational criteria for transfusion in preterm infants. PMID:8777681

  4. KIAA0101 is associated with human renal cell carcinoma proliferation and migration induced by erythropoietin

    PubMed Central

    Fan, Shengjun; Li, Xin; Tie, Lu; Pan, Yan; Li, Xuejun

    2016-01-01

    Erythropoietin (EPO) is a frequently prescribed anti-anemic drug for patients with advanced renal carcinoma. However, recent evidence from clinical studies suggested that EPO accelerated tumor progression and jeopardized the 5-year survival. Herein, we show, starting from the in silico microarray bioinformatics analysis, that activation of Erythropoietin signaling pathway enhanced renal clear carcinoma (RCC) progression. EPO accelerated the proliferative and migratory ability in 786-O and Caki-2 cells. Moreover, comparative proteomics expression profiling suggested that exogenous EPO stimulated RCC progression via up-regulation of KIAA0101 expression. Loss of KIAA0101 impeded the undesirable propensity of EPO in RCC. Finally, low expression of KIAA0101 was associated with the excellent prognosis and prognosticated a higher 5-year survival in human patients with renal carcinoma. Overall, KIAA0101 appears to be a key promoter of RCC malignancy induced by EPO, which provide mechanistic insights into KIAA0101 functions, and pave the road to develop new therapeutics for treatment of cancer-related and chemotherapy-induced anemia in patients with RCC. PMID:26575329

  5. Erythropoietin and IGF-1 signaling synchronize cell proliferation and maturation during erythropoiesis.

    PubMed

    Kadri, Zahra; Lefevre, Carine; Goupille, Olivier; Penglong, Tipparat; Granger-Locatelli, Marine; Fucharoen, Suthat; Maouche-Chretien, Leila; Leboulch, Philippe; Chretien, Stany

    2015-12-15

    Tight coordination of cell proliferation and differentiation is central to red blood cell formation. Erythropoietin controls the proliferation and survival of red blood cell precursors, while variations in GATA-1/FOG-1 complex composition and concentrations drive their maturation. However, clear evidence of cross-talk between molecular pathways is lacking. Here, we show that erythropoietin activates AKT, which phosphorylates GATA-1 at Ser310, thereby increasing GATA-1 affinity for FOG-1. In turn, FOG-1 displaces pRb/E2F-2 from GATA-1, ultimately releasing free, proproliferative E2F-2. Mice bearing a Gata-1(S310A) mutation suffer from fatal anemia when a compensatory pathway for E2F-2 production involving insulin-like growth factor-1 (IGF-1) signaling is simultaneously abolished. In the context of the GATA-1(V205G) mutation resulting in lethal anemia, we show that the Ser310 cannot be phosphorylated and that constitutive phosphorylation at this position restores partial erythroid differentiation. This study sheds light on the GATA-1 pathways that synchronize cell proliferation and differentiation for tissue homeostasis. PMID:26680303

  6. Erythropoietin therapy after allogeneic hematopoietic cell transplantation: a prospective, randomized trial.

    PubMed

    Jaspers, Aurélie; Baron, Frédéric; Willems, Evelyne; Seidel, Laurence; Hafraoui, Kaoutar; Vanstraelen, Gaetan; Bonnet, Christophe; Beguin, Yves

    2014-07-01

    We conducted a prospective randomized trial to assess hemoglobin (Hb) response to recombinant human erythropoietin (rhEPO) therapy after hematopoietic cell transplantation (HCT). Patients (N = 131) were randomized (1:1) between no treatment (control arm) or erythropoietin at 500 U/kg per week (EPO arm). Patients were also stratified into 3 cohorts: patients undergoing myeloablative HCT with rhEPO to start on day (D)28, patients given nonmyeloablative HCT (NMHCT) with rhEPO to start on D28, and patients also given NMHCT but with rhEPO to start on D0. The proportion of complete correctors (ie, Hb ≥13 g/dL) before D126 posttransplant was 8.1% in the control arm (median not reached) and 63.1% in the EPO arm (median, 90 days) (P < .001). Hb levels were higher and transfusion requirements decreased (P < .001) in the EPO arm, but not during the first month in the nonmyeloablative cohort starting rhEPO on D0. There was no difference in rates of thromboembolic events or other complications between the 2 arms. This is the first randomized trial to demonstrate that rhEPO therapy hastens erythroid recovery and decreases transfusion requirements when started one month after allogeneic HCT. There was no benefit to start rhEPO earlier after NMHCT.

  7. Erythropoietin and IGF-1 signaling synchronize cell proliferation and maturation during erythropoiesis

    PubMed Central

    Kadri, Zahra; Lefevre, Carine; Goupille, Olivier; Penglong, Tipparat; Granger-Locatelli, Marine; Fucharoen, Suthat; Maouche-Chretien, Leila; Leboulch, Philippe; Chretien, Stany

    2015-01-01

    Tight coordination of cell proliferation and differentiation is central to red blood cell formation. Erythropoietin controls the proliferation and survival of red blood cell precursors, while variations in GATA-1/FOG-1 complex composition and concentrations drive their maturation. However, clear evidence of cross-talk between molecular pathways is lacking. Here, we show that erythropoietin activates AKT, which phosphorylates GATA-1 at Ser310, thereby increasing GATA-1 affinity for FOG-1. In turn, FOG-1 displaces pRb/E2F-2 from GATA-1, ultimately releasing free, proproliferative E2F-2. Mice bearing a Gata-1S310A mutation suffer from fatal anemia when a compensatory pathway for E2F-2 production involving insulin-like growth factor-1 (IGF-1) signaling is simultaneously abolished. In the context of the GATA-1V205G mutation resulting in lethal anemia, we show that the Ser310 cannot be phosphorylated and that constitutive phosphorylation at this position restores partial erythroid differentiation. This study sheds light on the GATA-1 pathways that synchronize cell proliferation and differentiation for tissue homeostasis. PMID:26680303

  8. CNS Hypoxia Is More Pronounced in Murine Cerebral than Noncerebral Malaria and Is Reversed by Erythropoietin

    PubMed Central

    Hempel, Casper; Combes, Valery; Hunt, Nicholas Henry; Kurtzhals, Jørgen Anders Lindholm; Grau, Georges Emile Raymond

    2011-01-01

    Cerebral malaria (CM) is associated with high mortality and risk of sequelae, and development of adjunct therapies is hampered by limited knowledge of its pathogenesis. To assess the role of cerebral hypoxia, we used two experimental models of CM, Plasmodium berghei ANKA in CBA and C57BL/6 mice, and two models of malaria without neurologic signs, P. berghei K173 in CBA mice and P. berghei ANKA in BALB/c mice. Hypoxia was demonstrated in brain sections using intravenous pimonidazole and staining with hypoxia-inducible factor-1α–specific antibody. Cytopathic hypoxia was studied using poly (ADP-ribose) polymerase-1 (PARP-1) gene knockout mice. The effect of erythropoietin, an oxygen-sensitive cytokine that mediates protection against CM, on cerebral hypoxia was studied in C57BL/6 mice. Numerous hypoxic foci of neurons and glial cells were observed in mice with CM. Substantially fewer and smaller foci were observed in mice without CM, and hypoxia seemed to be confined to neuronal cell somas. PARP-1–deficient mice were not protected against CM, which argues against a role for cytopathic hypoxia. Erythropoietin therapy reversed the development of CM and substantially reduced the degree of neural hypoxia. These findings demonstrate cerebral hypoxia in malaria, strongly associated with cerebral dysfunction and a possible target for adjunctive therapy. PMID:21854739

  9. Recombinant human erythropoietin stimulates angiogenesis and healing of ischemic skin wounds.

    PubMed

    Buemi, Michele; Galeano, Mariarosaria; Sturiale, Alessio; Ientile, Riccardo; Crisafulli, Costantino; Parisi, Alessandra; Catania, MariaAntonietta; Calapai, Gioacchino; Impalà, Patrizia; Aloisi, Carmela; Squadrito, Francesco; Altavilla, Domenica; Bitto, Alessandra; Tuccari, Giovanni; Frisina, Nicola

    2004-08-01

    Wound healing in ischemic tissues such as flap margins due to inadequate blood supply is still a source of considerable morbidity in surgical practice. Adequate tissue perfusion is particularly important in wound healing. We investigated the effects of recombinant human erythropoietin (rHuEPO) on wound healing in an ischemic skin wound model. Sixty-three Sprague-Dawley rats were used. Normal incisional wound and H-shaped double flaps were used as the wound models. Animals were treated with rHuEPO (400 IU/kg) or its vehicle. Rats were killed on different days (3, 5, and 10 days after skin injury) and the wounded skin tissues were used for immunohistochemistry and for analysis of vascular endothelial growth factor content and collagen content. Tissue transglutaminase immunostaining of histological specimens was used as a vascular marker to determine the level of microvessel density. The results showed a higher level of vascular endothelial growth factor protein and an increased microvessel density in ischemic wounds with rHuEPO treatment than the normal incisional wounds and ischemic control wounds. Collagen content was higher in the incisional wounds and in the ischemic wounds with rHuEPO treatment compared with the ischemic control wounds. Our results suggest that erythropoietin may be an effective therapeutic approach in improving healing in ischemic skin wounds.

  10. Effects of ganglioside G(M1) and erythropoietin on spinal cord lesions in rats: functional and histological evaluations

    PubMed Central

    Marcon, Raphael Martus; Cristante, Alexandre Fogaça; de Barros Filho, Tarcísio Eloy Pessoa; Ferreira, Ricardo; dos Santos, Gustavo Bispo

    2016-01-01

    OBJECTIVE: To evaluate the functional and histological effects of ganglioside G(M1) and erythropoietin after experimental spinal cord contusion injury. METHODS: Fifty male Wistar rats underwent experimental spinal cord lesioning using an NYU-Impactor device and were randomly divided into the following groups, which received treatment intraperitoneally. The G(M1) group received ganglioside G(M1) (30 mg/kg); the erythropoietin group received erythropoietin (1000 IU/kg); the combined group received both drugs; and the saline group received saline (0.9%) as a control. A fifth group was the laminectomy group, in which the animals were subjected to laminectomy alone, without spinal lesioning or treatment. The animals were evaluated according to the Basso, Beattie and Bresnahan (BBB) scale, motor evoked potential recordings and, after euthanasia, histological analysis of spinal cord tissue. RESULTS: The erythropoietin group had higher BBB scores than the G(M1) group. The combined group had the highest BBB scores, and the saline group had the lowest BBB scores. No significant difference in latency was observed between the three groups that underwent spinal cord lesioning and intervention. However, the combined group showed a significantly higher signal amplitude than the other treatment groups or the saline group (p<0.01). Histological tissue analysis showed no significant difference between the groups. Axonal index was significantly enhanced in the combined group than any other intervention (p<0.01). CONCLUSION: G(M1) and erythropoietin exert therapeutic effects on axonal regeneration and electrophysiological and motor functions in rats subjected to experimental spinal cord lesioning and administering these two substances in combination potentiates their effects. PMID:27438570

  11. The effect of recombinant human erythropoietin (r-HuEPO) on left ventricular mass and left ventricular hemodynamics in hemodialysis patients.

    PubMed

    Minagawa, T; Hirano, T; Takada, N; Ishiguro, M; Ohtsuka, H

    1994-08-01

    The purpose of this study was to examine the effect of recombinant human erythropoietin (r-HuEPO) on left ventricular mass. Twenty-seven hemodialysis patients (13 men and 14 women) were given r-HuEPO for renal anemia. Blood pressure and heart rate were measured before and after the 16-week course of r-HuEPO, and at the same time echocardiography was performed to measure left ventricular dimensions and wall thickness. These measurements were used to calculate left ventricular volume, cardiac output (CO), and left ventricular mass (LVmass). Diastolic blood pressure (DBP) increased after administration of r-HuEPO (from 75.8 +/- 10.8 mmHg to 85.6 +/- 12.7 mmHg), but there was no change in systolic blood pressure (SBP) or heart rate. LVmass increased significantly in seven cases (from 194.7 +/- 40.0 g to 240.3 +/- 47.3 g). These cases, Group I, showed no decline in stroke volume (SV) or CO, and showed significant increases in SBP. In the remaining 20 cases, Group II, LVmass decreased or was unchanged. In this group SV and CO decreased, but there was no increase in SBP. We conclude that increases in LVmass may be associated with elevated systolic blood pressure and hypertrophy of the left ventricular wall, when hemodialysis patients with severe renal anemia are given r-HuEPO. PMID:7933670

  12. Apaf-1, Bcl-xL, cytochrome c, and caspase-9 form the critical elements for cerebral vascular protection by erythropoietin.

    PubMed

    Chong, Zhao Zhong; Kang, Jing-Qiong; Maiese, Kenneth

    2003-03-01

    Erythropoietin (EPO) plays a prominent role in the regulation of the hematopoietic system, but the potential function of this trophic factor as a cytoprotectant in the cerebral vascular system is not known. The authors examined the ability of EPO to modulate a series of death-related cellular pathways during free radical-induced injury in cerebral microvascular endothelial cells (ECs). Endothelial cell injury was evaluated by trypan blue, DNA fragmentation, membrane phosphatidylserine exposure, apoptotic protease-activating factor-1 (Apaf-1), and Bcl-XL expression, mitochondrial membrane potential, cytochrome c release, and cysteine protease activity. They show that constitutive EPO is present in ECs but is insufficient to prevent cellular injury. Signaling through the EPO receptor, however, remains biologically responsive to exogenous EPO administration to offer significant protection against nitric oxide-induced injury. Exogenous EPO maintains both genomic DNA integrity and cellular membrane asymmetry through parallel pathways that prevent the induction of Apaf-1 and preserve mitochondrial membrane potential in conjunction with enhanced Bcl-XL expression. Consistent with the modulation of Apaf-1 and the release of cytochrome c, EPO also inhibits the activation of caspase-9 and caspase-3-like activities. Identification of novel cytoprotective pathways used by EPO may serve as therapeutic targets for cerebral vascular disease.

  13. Erythropoietin administration alone or in combination with endurance training affects neither skeletal muscle morphology nor angiogenesis in healthy young men.

    PubMed

    Larsen, Mads S; Vissing, Kristian; Thams, Line; Sieljacks, Peter; Dalgas, Ulrik; Nellemann, Birgitte; Christensen, Britt

    2014-10-01

    The aim was to investigate the ability of an erythropoiesis-stimulating agent (ESA), alone or in combination with endurance training, to induce changes in human skeletal muscle fibre and vascular morphology. In a comparative study, 36 healthy untrained men were randomly dispersed into the following four groups: sedentary-placebo (SP, n = 9); sedentary-ESA (SE, n = 9); training-placebo (TP, n = 10); or training-ESA (TE, n = 8). The ESA or placebo was injected once weekly. Training consisted of progressive bicycling three times per week for 10 weeks. Before and after the intervention period, muscle biopsies and magnetic resonance images were collected from the thigh muscles, blood was collected, body composition measured and endurance exercise performance evaluated. The ESA treatment (SE and TE) led to elevated haematocrit, and both ESA treatment and training (SE, TP and TE) increased maximal O2 uptake. With regard to skeletal muscle morphology, TP alone exhibited increases in whole-muscle cross-sectional area and fibre diameter of all fibre types. Also exclusively for TP was an increase in type IIa fibres and a corresponding decrease in type IIx fibres. Furthermore, an overall training effect (TP and TE) was statistically demonstrated in whole-muscle cross-sectional area, muscle fibre diameter and type IIa and type IIx fibre distribution. With regard to muscle vascular morphology, TP and TE both promoted a rise in capillary to muscle fibre ratio, with no differences between the two groups. There were no effects of ESA treatment on any of the muscle morphological parameters. Despite the haematopoietic effects of ESA, we provide novel evidence that endurance training rather than ESA treatment induces adaptational changes in angiogenesis and muscle morphology.

  14. Monthly continuous erythropoietin receptor activator treatment maintains stable hemoglobin levels in routine clinical management of hemodialysis patients.

    PubMed

    Weinreich, Thomas; Leistikow, Frank; Hartmann, Hagen-Georg; Vollgraf, Günter; Dellanna, Frank

    2012-01-01

    Once-monthly administration of CERA, a continuous erythropoietin receptor activator, has shown equivalent efficacy to shorter-acting erythropoiesis-stimulating agents (ESAs) that require more frequent dosing, but data on routine use of once-monthly CERA in hemodialysis patients are lacking. Study on Efficacy, Safety and Applicability of Mircera (SESAM) was a prospective, multicenter, noninterventional trial with a duration of up to 9 months (month 0-5 "titration phase"; month 6-8 "evaluation phase") to test the stability of Hb control in hemodialysis patients under routine conditions. Patient selection, Hb targets and CERA dosing were at the discretion of the local nephrologist. 918 patients from 92 German nephrology centers were included. Ninety-three percent were on ESA treatment prior to study entry. The mean number of CERA dose changes during the study was 1.9 ± 1.9 per patient. Mean Hb level was 11.4 ± 1.2 g/dL at baseline and 11.7 ± 1.4 g/dL at the end of the 8-month study. During the evaluation phase (months 6-8), 15.6%, 40.3%, and 66.0% of patients had stable Hb (i.e., at least two values) in the ranges 11-12, 10-12, and 10-13 g/dL, respectively. The mean intra-individual fluctuation in Hb was 1.4 ± 0.7 g/dL during the study (0.5 ± 0.4 g/dL during the 3-month evaluation phase). More than 90% of patients, and > 80% of physicians, rated CERA therapy as "very good" or "good" throughout the study. Four patients (0.4%) discontinued prematurely due to adverse drug reactions. Once-monthly CERA therapy maintains stable Hb values with low intra-individual variability and few dose adaptations in hemodialysis patients when administered entirely according to local practice, and the regimen was well-tolerated.

  15. Administrative IT

    ERIC Educational Resources Information Center

    Grayson, Katherine, Ed.

    2006-01-01

    When it comes to Administrative IT solutions and processes, best practices range across the spectrum. Enterprise resource planning (ERP), student information systems (SIS), and tech support are prominent and continuing areas of focus. But widespread change can also be accomplished via the implementation of campuswide document imaging and sharing,…

  16. Database Administrator

    ERIC Educational Resources Information Center

    Moore, Pam

    2010-01-01

    The Internet and electronic commerce (e-commerce) generate lots of data. Data must be stored, organized, and managed. Database administrators, or DBAs, work with database software to find ways to do this. They identify user needs, set up computer databases, and test systems. They ensure that systems perform as they should and add people to the…

  17. ADMINISTRATIVE CLIMATE.

    ERIC Educational Resources Information Center

    BRUCE, ROBERT L.; CARTER, G.L., JR.

    IN THE COOPERATIVE EXTENSION SERVICE, STYLES OF LEADERSHIP PROFOUNDLY AFFECT THE QUALITY OF THE SERVICE RENDERED. ACCORDINGLY, MAJOR INFLUENCES ON ADMINISTRATIVE CLIMATE AND EMPLOYEE PRODUCTIVITY ARE EXAMINED IN ESSAYS ON (1) SOURCES OF JOB SATISFACTION AND DISSATISFACTION, (2) MOTIVATIONAL THEORIES BASED ON JOB-RELATED SATISFACTIONS AND NEEDS,…

  18. Engineering Administration.

    ERIC Educational Resources Information Center

    Naval Personnel Program Support Activity, Washington, DC.

    This book is intended to acquaint naval engineering officers with their duties in the engineering department. Standard shipboard organizations are analyzed in connection with personnel assignments, division operations, and watch systems. Detailed descriptions are included for the administration of directives, ship's bills, damage control, training…

  19. Erythropoietin-regulated oxidative stress negatively affects enucleation during terminal erythropoiesis.

    PubMed

    Zhao, Baobing; Mei, Yang; Yang, Jing; Ji, Peng

    2016-10-01

    Differentiating erythroblasts are exposed to an oxidative environment. The dynamics of oxidative status during terminal erythropoiesis and how they affect cell differentiation in response to erythropoietin (Epo) are unclear. Here, we show that Epo induces reactive oxygen species (ROS) production in the early stages of terminal erythropoiesis. The levels of ROS correlate with CD71 surface expression and the uptake of iron and transferrin. ROS decreases in the late stages of terminal erythropoiesis, when the cells are preparing for enucleation. Consistently, treatment of erythroblasts with a low dose (5 mM) of N-acetyl-cysteine (NAC), a ROS scavenger, promotes enucleation. However, a high dose (20 mM) of NAC leads to significant cell death. Our study reveals an important function of Epo in regulating the dynamics of oxidative status and enucleation. PMID:27364565

  20. [Pharma-clinics. Doping with erythropoietin or the misuse of therapeutic advances].

    PubMed

    Scheen, A J

    1998-08-01

    Sport performance depends on the capacity of oxygen transport to the exercising skeletal muscles. Performance may be increased thanks to an haematocrit augmentation, either by training in high altitude, using blood transfusions, or injecting erythropoietin (Epo). Since the synthesis of Epo by bioengineering, doping with recombinant human Epo has become popular in sports in general, and in cycling in particular. This new kind of doping might have been responsible for several sudden deaths in young athletes, essentially in racing cyclists. Until now, such a doping is hardly to be detected. However, it should deserve careful consideration from both public health authorities and sports communities in order to stop what could become a real plague.

  1. Erythropoietin-regulated oxidative stress negatively affects enucleation during terminal erythropoiesis.

    PubMed

    Zhao, Baobing; Mei, Yang; Yang, Jing; Ji, Peng

    2016-10-01

    Differentiating erythroblasts are exposed to an oxidative environment. The dynamics of oxidative status during terminal erythropoiesis and how they affect cell differentiation in response to erythropoietin (Epo) are unclear. Here, we show that Epo induces reactive oxygen species (ROS) production in the early stages of terminal erythropoiesis. The levels of ROS correlate with CD71 surface expression and the uptake of iron and transferrin. ROS decreases in the late stages of terminal erythropoiesis, when the cells are preparing for enucleation. Consistently, treatment of erythroblasts with a low dose (5 mM) of N-acetyl-cysteine (NAC), a ROS scavenger, promotes enucleation. However, a high dose (20 mM) of NAC leads to significant cell death. Our study reveals an important function of Epo in regulating the dynamics of oxidative status and enucleation.

  2. Is erythropoietin a worthy candidate for traumatic brain injury or are we heading the wrong way?

    PubMed Central

    Grasso, Giovanni; Alafaci, Concetta; Ghezzi, Pietro

    2016-01-01

    Traumatic brain injury (TBI) is a leading cause of death and disability in the modern society. Although primary prevention is the only strategy that can counteract the primary brain damage, numerous preclinical studies have been accumulated in order to find therapeutic strategies against the secondary damage. In this scenario erythropoietin (EPO) has been shown to be a promising candidate as neuroprotective agent. A recent clinical trial, however, has shown that EPO has not an overall effect on outcomes following TBI thus renewing old concerns.  However, the results of a prespecified sensitivity analysis indicate that the effect of EPO on mortality remains still unclear. In the light of these observations, further investigations are needed to resolve doubts on EPO effectiveness in order to provide a more solid base for tailoring conclusive clinical trials. PMID:27239280

  3. The neuroprotective effect of erythropoietin on experimental Parkinson model in rats.

    PubMed

    Erbaş, Oytun; Çınar, Bilge Piri; Solmaz, Volkan; Çavuşoğlu, Türker; Ateş, Utku

    2015-02-01

    Dopaminergic neuronal loss in Parkinson's disease (PD) results from oxidative stress, neuroinflammation and excitotoxicity. Because erythropoietin (EPO) has been shown to have antioxidant, anti-inflammatory and neuroprotective effects in many previous studies, present study was designed to evaluate the effect of EPO on rotenone-induced dopaminergic neuronal loss. The rats in which PD was induced by stereotaxical infusion of rotenone showed increased MDA and TNF-alpha levels and decreased HVA levels. On the other hand, EPO treatment resulted in markedly decreased MDA and TNF-alpha levels and increased HVA levels. EPO treatment in rotenone-infusion group resulted in improvement of striatal neurodegeneration and a significant increase in decreased total number of neurons and immunohistochemical TH positive neurons. Results of the present study demonstrate the neuroprotective, anti-inflammatory and antioxidant effects of EPO in a rotenone-induced neurodegenerative animal model. PMID:25464888

  4. Neuroprotection with erythropoietin in preterm and/or low birth weight infants.

    PubMed

    Zhang, Jie; Wang, Qiuxia; Xiang, Hong; Xin, Yue; Chang, Ming; Lu, Hongyan

    2014-08-01

    Neonatal brain injury caused by extreme prematurity remains a great challenge for prevention. Erythropoietin (EPO) has shown neuroprotective effects in a series of neonatal experimental models and recent clinical trials of premature infants. In this meta-analysis of seven clinical trials, EPO was associated with a highly reproducible reduction in the risk of neurodevelopmental disability in preterm infants. However, there was no difference in the risk for morbidity, cerebral palsy, visual deficit, severe hearing deficit, necrotizing enterocolitis, intracranial hemorrhage and patent ductus arteriosus. The use of EPO, to some extent, is associated with reduction in neurodevelopmental disability in preterm infants. More double blind randomized controlled trials are needed to establish the best therapeutic approach for neuroprotection in preterm infants. PMID:24650681

  5. [Use of recombinant human erythropoietin preparations as a blood-saving method in neurosurgery].

    PubMed

    Gromova, V V; Imaev, A A; Lubnin, A Iu

    2010-01-01

    The paper gives the results of the first clinical use of recombinant human erythropoietin (rhEPO) within the blood-saving program in neurosurgical patients with preoperative anemia of various genesis and predictable massive intraoperative blood loss. A course of effective therapy with rhEPO is shown to increase hemoglobin and packed cell volume rather rapidly in all patients prior to surgery. This made it possible to effectively apply other blood-saving procedures (isovolemic hemodilution and instrumental reinfusion of washed red blood cells) and to reduce the volume required for the use of donor transfused media in these patients. Various clinical aspects of the use of rhEPO in these patients are discussed.

  6. Chemical synthesis of erythropoietin glycoforms for insights into the relationship between glycosylation pattern and bioactivity.

    PubMed

    Murakami, Masumi; Kiuchi, Tatsuto; Nishihara, Mika; Tezuka, Katsunari; Okamoto, Ryo; Izumi, Masayuki; Kajihara, Yasuhiro

    2016-01-01

    The role of sialyloligosaccharides on the surface of secreted glycoproteins is still unclear because of the difficulty in the preparation of sialylglycoproteins in a homogeneous form. We selected erythropoietin (EPO) as a target molecule and designed an efficient synthetic strategy for the chemical synthesis of a homogeneous form of five EPO glycoforms varying in glycosylation position and the number of human-type biantennary sialyloligosaccharides. A segment coupling strategy performed by native chemical ligation using six peptide segments including glycopeptides yielded homogeneous EPO glycopeptides, and folding experiments of these glycopeptides afforded the correctly folded EPO glycoforms. In an in vivo erythropoiesis assay in mice, all of the EPO glycoforms displayed biological activity, in particular the EPO bearing three sialyloligosaccharides, which exhibited the highest activity. Furthermore, we observed that the hydrophilicity and biological activity of the EPO glycoforms varied depending on the glycosylation pattern. This knowledge will pave the way for the development of homogeneous biologics by chemical synthesis.

  7. Chemical synthesis of erythropoietin glycoforms for insights into the relationship between glycosylation pattern and bioactivity

    PubMed Central

    Murakami, Masumi; Kiuchi, Tatsuto; Nishihara, Mika; Tezuka, Katsunari; Okamoto, Ryo; Izumi, Masayuki; Kajihara, Yasuhiro

    2016-01-01

    The role of sialyloligosaccharides on the surface of secreted glycoproteins is still unclear because of the difficulty in the preparation of sialylglycoproteins in a homogeneous form. We selected erythropoietin (EPO) as a target molecule and designed an efficient synthetic strategy for the chemical synthesis of a homogeneous form of five EPO glycoforms varying in glycosylation position and the number of human-type biantennary sialyloligosaccharides. A segment coupling strategy performed by native chemical ligation using six peptide segments including glycopeptides yielded homogeneous EPO glycopeptides, and folding experiments of these glycopeptides afforded the correctly folded EPO glycoforms. In an in vivo erythropoiesis assay in mice, all of the EPO glycoforms displayed biological activity, in particular the EPO bearing three sialyloligosaccharides, which exhibited the highest activity. Furthermore, we observed that the hydrophilicity and biological activity of the EPO glycoforms varied depending on the glycosylation pattern. This knowledge will pave the way for the development of homogeneous biologics by chemical synthesis. PMID:26824070

  8. Role of cytochrome P sub 450 in the control of the production of erythropoietin

    SciTech Connect

    Fandrey, J.; Seydel, F.P.; Siegers, C.P.; Jelkmann, W. )

    1990-01-01

    Effects of agents affecting cytochrome P{sub 450} were studied on the production of erythropoietin (Epo) in cultures of the human hepatoma cell line HepG2. Epo was measured by radioimmunoassay of the culture media after 24 h of incubation. The addition of phenobarbital or 3-methylcholanthrene, which induce cytochrome P{sub 450}, significantly enhanced the formation of Epo. Likewise, the thyroid hormones T{sub 3} and T{sub 4} stimulated the rate of the production of Epo. On the other hand, the formation of Epo was lowered following the addition of diethyl-dithiocarbamate or cysteamine chloride, which inhibit cytochrome P{sub 450}. These findings support the idea that O{sub 2} sensitive hemoproteins of the microsomal mixed-functional oxidases play a role in the control of the synthesis of Epo.

  9. Validation of model virus removal and inactivation capacity of an erythropoietin purification process.

    PubMed

    Pérez, Mayté; Rodríguez, Elias; Rodríguez, María; Paez, Rolando; Ruibal, Ignacio; Noa, Enrique; García, Osnel; Moya, Galina; Martínez, Mayda; Marcelo, José; Martínez, Anazuria; Dubal, Marta; Navea, Leonor; Valdés, Rodolfo

    2011-11-01

    Human erythropoietin (hEpo) production requires mammalian cells able to make complex post-translational modifications to guaranty its biological activity. As mammalian cell can be reservoir of pathogenic viruses and several animal origin components are usually used in the cultivation of mammalian cells, hEpo contamination with viruses is something of great concern. As consequence, this study investigated the viral removal and inactivation capacity of a recombinant-hEpo (rec-hEpo) purification process. Canine parvovirus, Human poliovirus type-2, Bovine viral diarrhea virus and Human immunodeficiency virus type-1 were used for measuring process viral removal and inactivation capacities. In conclusion, this study corroborated that the assessed rec-hEpo purification process has enough capacity (5.0-19.4 Logs) for removing and inactivating these model viruses and sodium hydroxide demonstrated to be a robust sanitization solution for chromatography columns (5.0 (PV-2)-6.7 (CPV) Logs).

  10. Widespread Expression of Erythropoietin Receptor in Brain and Its Induction by Injury

    PubMed Central

    Ott, Christoph; Martens, Henrik; Hassouna, Imam; Oliveira, Bárbara; Erck, Christian; Zafeiriou, Maria-Patapia; Peteri, Ulla-Kaisa; Hesse, Dörte; Gerhart, Simone; Altas, Bekir; Kolbow, Tekla; Stadler, Herbert; Kawabe, Hiroshi; Zimmermann, Wolfram-Hubertus; Nave, Klaus-Armin; Schulz-Schaeffer, Walter; Jahn, Olaf; Ehrenreich, Hannelore

    2015-01-01

    Erythropoietin (EPO) exerts potent neuroprotective, neuroregenerative and procognitive functions. However, unequivocal demonstration of erythropoietin receptor (EPOR) expression in brain cells has remained difficult since previously available anti-EPOR antibodies (EPOR-AB) were unspecific. We report here a new, highly specific, polyclonal rabbit EPOR-AB directed against different epitopes in the cytoplasmic tail of human and murine EPOR and its characterization by mass spectrometric analysis of immuno-precipitated endogenous EPOR, Western blotting, immunostaining and flow cytometry. Among others, we applied genetic strategies including overexpression, Lentivirus-mediated conditional knockout of EpoR and tagged proteins, both on cultured cells and tissue sections, as well as intracortical implantation of EPOR-transduced cells to verify specificity. We show examples of EPOR expression in neurons, oligodendroglia, astrocytes and microglia. Employing this new EPOR-AB with double-labeling strategies, we demonstrate membrane expression of EPOR as well as its localization in intracellular compartments such as the Golgi apparatus. Moreover, we show injury-induced expression of EPOR. In mice, a stereotactically applied stab wound to the motor cortex leads to distinct EpoR expression by reactive GFAP-expressing cells in the lesion vicinity. In a patient suffering from epilepsy, neurons and oligodendrocytes of the hippocampus strongly express EPOR. To conclude, this new analytical tool will allow neuroscientists to pinpoint EPOR expression in cells of the nervous system and to better understand its role in healthy conditions, including brain development, as well as under pathological circumstances, such as upregulation upon distress and injury. PMID:26349059

  11. Erythropoietin Therapy, Hemoglobin Targets, and Quality of Life in Healthy Hemodialysis Patients: A Randomized Trial

    PubMed Central

    Foley, Robert N.; Curtis, Bryan M.; Parfrey, Patrick S.

    2009-01-01

    Background and objectives: The effects of different hemoglobin targets when using erythropoiesis-stimulating agents on quality of life are somewhat controversial, and predictors of change in quality of life in endstage renal disease have not been well characterized. Design, setting, participants, & measurements: Five hundred ninety-six incident hemodialysis patients without symptomatic cardiac disease were randomly assigned to hemoglobin targets of 9.5 to 11.5 g/dl or 13.5 to 14.5 g/dl for 96 weeks, using epoetin_alfa as primary therapy. Patients and attending physicians were masked to treatment assignment. Quality of life, a secondary outcome, was prospectively recorded using the Kidney Disease Quality of Life (KDQoL) questionnaire at weeks 0, 24, 36, 48, 60, 72, 84, and 96, with prespecified outcomes being fatigue and quality of social interaction. Results: The mean age and prior duration of dialysis therapy of the study population were 50.8 and 0.8 yr. Mortality was low, reflecting the relatively healthy group enrolled. Of 20 domains within the KDQoL only the prespecified domain of fatigue showed significant change over time between the two groups. Improvement in fatigue scores in the high-target group ranged from 3.2 to 7.9 over time (P = 0.007) compared with change in the low-target group. Higher body mass index and lower erythropoietin dose at baseline were independent predictors of improvement in multiple KDQoL domains. Conclusions: In relatively healthy hemodialysis patients, normal hemoglobin targets may have beneficial effects on fatigue. Improvement in multiple domains of quality of life is associated with higher body mass index and lower erythropoietin requirements. PMID:19339412

  12. Erythropoietin is involved in the angiogenic potential of bone marrow macrophages in multiple myeloma.

    PubMed

    De Luisi, Annunziata; Binetti, Laura; Ria, Roberto; Ruggieri, Simona; Berardi, Simona; Catacchio, Ivana; Racanelli, Vito; Pavone, Vincenzo; Rossini, Bernardo; Vacca, Angelo; Ribatti, Domenico

    2013-10-01

    Erythropoietin (Epo) is the crucial cytokine regulator of red blood cell production, and recombinant human erythropoietin (rHuEpo) is widely used in clinical practice for the treatment of anemia, primarily in kidney disease and in cancer. Increasing evidence suggests several biological roles for Epo and its receptor, Epo-R, unrelated to erythropoiesis, including angiogenesis. Epo-R has been found expressed in various non-haematopoietic cells and tissues, and in cancer cells. Here, we detected the expression of Epo-R in bone marrow-derived macrophages (BMMAs) from multiple myeloma (MM) and monoclonal gammopathy of undetermined significance (MGUS) patients and assessed whether Epo/Epo-R axis plays a role in MM macrophage-mediated angiogenesis. We found that Epo-R is over-expressed in BMMAs from MM patients with active disease compared to MGUS patients. The treatment of BMMAs with rHuEpo significantly increased the expression and secretion of key pro-angiogenic mediators, such as vascular endothelial growth factor, hepatocyte growth factor and monocyte chemotactic protein (MCP-1/CCL-2), through activation of JAK2/STAT5 and PI3 K/Akt pathways. In addition, the conditioned media harvested from rHuEpo-treated BMMAs enhanced bone marrow-derived endothelial cell migration and capillary morphogenesis in vitro, and induced angiogenesis in the chorioallantoic membrane of chick embryos in vivo. Furthermore, we found an increase in the circulating levels of several pro-angiogenic cytokines in serum of MM patients with anemia under treatment with Epo. Our findings highlight the direct effect of rHuEpo on macrophage-mediated production of pro-angiogenic factors, suggesting that Epo/Epo-R pathway may be involved in the regulation of angiogenic response occurring in MM.

  13. Mapping the Homodimer Interface of an Optimized, Artificial, Transmembrane Protein Activator of the Human Erythropoietin Receptor

    PubMed Central

    Bears, Zachary; Barrera, Francisco N.; Alonso, Miriam; Engelman, Donald M.; DiMaio, Daniel

    2014-01-01

    Transmembrane proteins constitute a large fraction of cellular proteins, and specific interactions involving membrane-spanning protein segments play an important role in protein oligomerization, folding, and function. We previously isolated an artificial, dimeric, 44-amino acid transmembrane protein that activates the human erythropoietin receptor (hEPOR) in trans. This artificial protein supports limited erythroid differentiation of primary human hematopoietic progenitor cells in vitro, even though it does not resemble erythropoietin, the natural ligand of this receptor. Here, we used a directed-evolution approach to explore the structural basis for the ability of transmembrane proteins to activate the hEPOR. A library that expresses thousands of mutants of the transmembrane activator was screened for variants that were more active than the original isolate at inducing growth factor independence in mouse cells expressing the hEPOR. The most active mutant, EBC5-16, supports erythroid differentiation in human cells with activity approaching that of EPO, as assessed by cell-surface expression of glycophorin A, a late-stage marker of erythroid differentiation. EBC5-16 contains a single isoleucine to serine substitution at position 25, which increases its ability to form dimers. Genetic studies confirmed the importance of dimerization for activity and identified the residues constituting the homodimer interface of EBC5-16. The interface requires a GxxxG dimer packing motif and a small amino acid at position 25 for maximal activity, implying that tight packing of the EBC5-16 dimer is a crucial determinant of activity. These experiments identified an artificial protein that causes robust activation of its target in a natural host cell, demonstrated the importance of dimerization of this protein for engagement of the hEPOR, and provided the framework for future structure-function studies of this novel mechanism of receptor activation. PMID:24788775

  14. Erythropoietin is involved in the angiogenic potential of bone marrow macrophages in multiple myeloma.

    PubMed

    De Luisi, Annunziata; Binetti, Laura; Ria, Roberto; Ruggieri, Simona; Berardi, Simona; Catacchio, Ivana; Racanelli, Vito; Pavone, Vincenzo; Rossini, Bernardo; Vacca, Angelo; Ribatti, Domenico

    2013-10-01

    Erythropoietin (Epo) is the crucial cytokine regulator of red blood cell production, and recombinant human erythropoietin (rHuEpo) is widely used in clinical practice for the treatment of anemia, primarily in kidney disease and in cancer. Increasing evidence suggests several biological roles for Epo and its receptor, Epo-R, unrelated to erythropoiesis, including angiogenesis. Epo-R has been found expressed in various non-haematopoietic cells and tissues, and in cancer cells. Here, we detected the expression of Epo-R in bone marrow-derived macrophages (BMMAs) from multiple myeloma (MM) and monoclonal gammopathy of undetermined significance (MGUS) patients and assessed whether Epo/Epo-R axis plays a role in MM macrophage-mediated angiogenesis. We found that Epo-R is over-expressed in BMMAs from MM patients with active disease compared to MGUS patients. The treatment of BMMAs with rHuEpo significantly increased the expression and secretion of key pro-angiogenic mediators, such as vascular endothelial growth factor, hepatocyte growth factor and monocyte chemotactic protein (MCP-1/CCL-2), through activation of JAK2/STAT5 and PI3 K/Akt pathways. In addition, the conditioned media harvested from rHuEpo-treated BMMAs enhanced bone marrow-derived endothelial cell migration and capillary morphogenesis in vitro, and induced angiogenesis in the chorioallantoic membrane of chick embryos in vivo. Furthermore, we found an increase in the circulating levels of several pro-angiogenic cytokines in serum of MM patients with anemia under treatment with Epo. Our findings highlight the direct effect of rHuEpo on macrophage-mediated production of pro-angiogenic factors, suggesting that Epo/Epo-R pathway may be involved in the regulation of angiogenic response occurring in MM. PMID:23881169

  15. Erythropoietin prevents sepsis-related acute kidney injury in rats by inhibiting NF-κB and upregulating endothelial nitric oxide synthase.

    PubMed

    Souza, Ana Carolina C Pessoa de; Volpini, Rildo A; Shimizu, Maria Heloísa; Sanches, Talita Rojas; Camara, Niels Olsen Saraiva; Semedo, Patrícia; Rodrigues, Camila Eleutério; Seguro, Antonio Carlos; Andrade, Lúcia

    2012-04-15

    The pathophysiology of sepsis involves complex cytokine and inflammatory mediator networks, a mechanism to which NF-κB activation is central. Downregulation of endothelial nitric oxide synthase (eNOS) contributes to sepsis-induced endothelial dysfunction. Erythropoietin (EPO) has emerged as a major tissue-protective cytokine in the setting of stress. We investigated the role of EPO in sepsis-related acute kidney injury using a cecal ligation and puncture (CLP) model. Wistar rats were divided into three primary groups: control (sham-operated); CLP; and CLP+EPO. EPO (4,000 IU/kg body wt ip) was administered 24 and 1 h before CLP. Another group of rats received N-nitro-l-arginine methyl ester (l-NAME) simultaneously with EPO administration (CLP+EPO+l-NAME). A fifth group (CLP+EPOtreat) received EPO at 1 and 4 h after CLP. At 48 h postprocedure, CLP+EPO rats presented significantly higher inulin clearance than did CLP and CLP+EPO+l-NAME rats; hematocrit levels, mean arterial pressure, and metabolic balance remained unchanged in the CLP+EPO rats; and inulin clearance was significantly higher in CLP+EPOtreat rats than in CLP rats. At 48 h after CLP, creatinine clearance was significantly higher in the CLP+EPO rats than in the CLP rats. In renal tissue, pre-CLP EPO administration prevented the sepsis-induced increase in macrophage infiltration, as well as preserving eNOS expression, EPO receptor (EpoR) expression, IKK-α activation, NF-κB activation, and inflammatory cytokine levels, thereby increasing survival. We conclude that this protection, which appears to be dependent on EpoR activation and on eNOS expression, is attributable, in part, to inhibition of the inflammatory response via NF-κB downregulation.

  16. Multiple Doses of Erythropoietin Impair Liver Regeneration by Increasing TNF-α, the Bax to Bcl-xL Ratio and Apoptotic Cell Death

    PubMed Central

    Cantré, Daniel; Abshagen, Kerstin; Menger, Michael D.; Vollmar, Brigitte

    2008-01-01

    Background Liver resection and the use of small-for-size grafts are restricted by the necessity to provide a sufficient amount of functional liver mass. Only few promising strategies to maximize liver regeneration are available. Apart from its erythropoiesis-stimulating effect, erythropoietin (EPO) has meanwhile been recognized as mitogenic, tissue-protective, and anti-apoptotic pleiotropic cytokine. Thus, EPO may support regeneration of hepatic tissue. Methodology Rats undergoing 68% hepatectomy received daily either high dose (5000 IU/kg bw iv) or low dose (500 IU/kg bw iv) recombinant human EPO or equal amounts of physiologic saline. Parameters of liver regeneration and hepatocellular apoptosis were assessed at 24 h, 48 h and 5 d after resection. In addition, red blood cell count, hematocrit and serum EPO levels as well as plasma concentrations of TNF-α and IL-6 were evaluated. Further, hepatic Bcl-xL and Bax protein expression were analyzed by Western blot. Principal Findings Administration of EPO significantly reduced the expression of PCNA at 24 h followed by a significant decrease in restitution of liver mass at day 5 after partial hepatectomy. EPO increased TNF-α levels and shifted the Bcl-xL to Bax ratio towards the pro-apoptotic Bax resulting in significantly increased hepatocellular apoptosis. Conclusions Multiple doses of EPO after partial hepatectomy increase hepatocellular apoptosis and impair liver regeneration in rats. Thus, careful consideration should be made in pre- and post-operative recombinant human EPO administration in the setting of liver resection and transplantation. PMID:19079544

  17. Effects of cobalt on haem proteins of erythropoietin-producing HepG2 cells in multicellular spheroid culture.

    PubMed

    Görlach, A; Fandrey, J; Holtermann, G; Acker, H

    1994-07-11

    The hypoxia-induced increase of spectrophotometrically measured light absorption at 560 nm, considered as reduced cytochrome b, in HepG2 cells is diminished after exposure to cobalt chloride (50 or 100 microM) for 18-36 h. The redox state of cytochrome c and cytochrome aa3, however, remains stable, indicating a particular affinity of cytochrome b for cobalt. Erythropoietin production of HepG2 cells increases after application of cobalt chloride, whereas H2O2 production, as measured by the dihydrorhodamine technique, decreases. It is concluded that cobalt stimulates a signal cascade with cytochrome b as receptor and H2O2 as second messenger for regulating erythropoietin production.

  18. Effects of an 8-weeks erythropoietin treatment on mitochondrial and whole body fat oxidation capacity during exercise in healthy males.

    PubMed

    Guadalupe-Grau, Amelia; Plenge, Ulla; Helbo, Signe; Kristensen, Marianne; Andersen, Peter Riis; Fago, Angela; Belhage, Bo; Dela, Flemming; Helge, Jørn Wulff

    2015-01-01

    The present investigation was performed to elucidate if the non-erythropoietic ergogenic effect of a recombinant erythropoietin treatment results in an impact on skeletal muscle mitochondrial and whole body fatty acid oxidation capacity during exercise, myoglobin concentration and angiogenesis. Recombinant erythropoietin was administered by subcutaneous injections (5000 IU) in six healthy male volunteers (aged 21 ± 2 years; fat mass 18.5 ± 2.3%) over 8 weeks. The participants performed two graded cycle ergometer exercise tests before and after the intervention where VO2max and maximal fat oxidation were measured. Biopsies of the vastus lateralis muscle were obtained before and after the intervention. Recombinant erythropoietin treatment increased mitochondrial O2 flux during ADP stimulated state 3 respiration in the presence of complex I and II substrates (malate, glutamate, pyruvate, succinate) with additional electron input from β-oxidation (octanoylcarnitine) (from 60 ± 13 to 87 ± 24 pmol · s(-1) · mg(-1) P < 0.01). β-hydroxy-acyl-CoA-dehydrogenase activity was higher after treatment (P < 0.05), whereas citrate synthase activity also tended to increase (P = 0.06). Total myoglobin increased by 16.5% (P < 0.05). Capillaries per muscle area tended to increase (P = 0.07), whereas capillaries per fibre as well as the total expression of vascular endothelial growth factor remained unchanged. Whole body maximal fat oxidation was not increased after treatment. Eight weeks of recombinant erythropoietin treatment increases mitochondrial fatty acid oxidation capacity and myoglobin concentration without any effect on whole body maximal fat oxidation. PMID:25259652

  19. Treatment of optic neuritis with erythropoietin (TONE): a randomised, double-blind, placebo-controlled trial—study protocol

    PubMed Central

    Diem, Ricarda; Molnar, Fanni; Beisse, Flemming; Gross, Nikolai; Drüschler, Katharina; Heinrich, Sven P; Joachimsen, Lutz; Rauer, Sebastian; Pielen, Amelie; Sühs, Kurt-Wolfram; Linker, Ralf Andreas; Huchzermeyer, Cord; Albrecht, Philipp; Hassenstein, Andrea; Aktas, Orhan; Guthoff, Tanja; Tonagel, Felix; Kernstock, Christoph; Hartmann, Kathrin; Kümpfel, Tania; Hein, Katharina; van Oterendorp, Christian; Grotejohann, Birgit; Ihorst, Gabriele; Maurer, Julia; Müller, Matthias; Volkmann, Martin; Wildemann, Brigitte; Platten, Michael; Wick, Wolfgang; Heesen, Christoph; Schiefer, Ulrich; Wolf, Sebastian; Lagrèze, Wolf A

    2016-01-01

    Introduction Optic neuritis leads to degeneration of retinal ganglion cells whose axons form the optic nerve. The standard treatment is a methylprednisolone pulse therapy. This treatment slightly shortens the time of recovery but does not prevent neurodegeneration and persistent visual impairment. In a phase II trial performed in preparation of this study, we have shown that erythropoietin protects global retinal nerve fibre layer thickness (RNFLT-G) in acute optic neuritis; however, the preparatory trial was not powered to show effects on visual function. Methods and analysis Treatment of Optic Neuritis with Erythropoietin (TONE) is a national, randomised, double-blind, placebo-controlled, multicentre trial with two parallel arms. The primary objective is to determine the efficacy of erythropoietin compared to placebo given add-on to methylprednisolone as assessed by measurements of RNFLT-G and low-contrast visual acuity in the affected eye 6 months after randomisation. Inclusion criteria are a first episode of optic neuritis with decreased visual acuity to ≤0.5 (decimal system) and an onset of symptoms within 10 days prior to inclusion. The most important exclusion criteria are history of optic neuritis or multiple sclerosis or any ocular disease (affected or non-affected eye), significant hyperopia, myopia or astigmatism, elevated blood pressure, thrombotic events or malignancy. After randomisation, patients either receive 33 000 international units human recombinant erythropoietin intravenously for 3 consecutive days or placebo (0.9% saline) administered intravenously. With an estimated power of 80%, the calculated sample size is 100 patients. The trial started in September 2014 with a planned recruitment period of 30 months. Ethics and dissemination TONE has been approved by the Central Ethics Commission in Freiburg (194/14) and the German Federal Institute for Drugs and Medical Devices (61-3910-4039831). It complies with the Declaration of Helsinki

  20. Erythropoietin test

    MedlinePlus

    ... kidney. These cells release more EPO when blood oxygen level is low. How the Test is Performed A blood sample is needed. How ... your doctor about the meaning of your specific test result. What ... in response to an event such as low blood oxygen level. The condition may occur at high altitudes ...

  1. Erythropoietin inhibits gamma-irradiation-induced apoptosis by upregulation of Bcl-2 and decreasing the activation of caspase 3 in human UT-7/erythropoietin cell line.

    PubMed

    Liu, Yuan-Yuan; She, Zhen-Jue; Yao, Ming-Hui

    2010-05-01

    1. Erythropoietin (EPO) can reverse radiotherapy-induced anaemia by stimulating bone marrow cells to produce erythrocytes. However, there are limited studies that address the mechanisms by which EPO exerts its beneficial effects in radiotherapy-induced anaemia. In the present study, we used a human bone marrow-derived EPO-dependent leukaemia cell line UT-7/EPO that progressed further in erythroid development to evaluate the anti-apoptotic effects of EPO on irradiated human erythroid progenitor. 2. The UT-7/EPO cells exposed to gamma-irradiation were cultured in the presence or absence of EPO at a concentration of 7 U/mL. The cell viability, cell apoptosis and the expression of apoptosis-related proteins Bcl-2, Bax and caspase 3 were examined. 3. The results showed that EPO protected the viability of human UT-7/EPO cells exposed to gamma-irradiation. EPO significantly inhibited gamma-irradiation-induced apoptosis in human UT-7/EPO cells: a significant decrease in the percentage of apoptotic cells was observed (62, 69 and 62% at 24, 48 and 72 h, respectively). Furthermore, EPO significantly increased the expression of Bcl-2 protein and the relative Bcl-2/Bax ratio, and decreased the activation of caspase 3 and formation of the p17 and p12 cleavage in similar conditions. 4. In conclusion, EPO exerts anti-apoptotic effects on irradiated human UT-7/EPO cells through upregulation of Bcl-2 protein and the relative Bcl-2/Bax ratio, and by decreasing the activation of caspase 3. These findings may contribute to our understanding of the beneficial function of EPO in radiotherapy-induced anaemia.

  2. A simple three-step method for design and affinity testing of new antisense peptides: an example of erythropoietin.

    PubMed

    Štambuk, Nikola; Manojlović, Zoran; Turčić, Petra; Martinić, Roko; Konjevoda, Paško; Weitner, Tin; Wardega, Piotr; Gabričević, Mario

    2014-01-01

    Antisense peptide technology is a valuable tool for deriving new biologically active molecules and performing peptide-receptor modulation. It is based on the fact that peptides specified by the complementary (antisense) nucleotide sequences often bind to each other with a higher specificity and efficacy. We tested the validity of this concept on the example of human erythropoietin, a well-characterized and pharmacologically relevant hematopoietic growth factor. The purpose of the work was to present and test simple and efficient three-step procedure for the design of an antisense peptide targeting receptor-binding site of human erythropoietin. Firstly, we selected the carboxyl-terminal receptor binding region of the molecule (epitope) as a template for the antisense peptide modeling; Secondly, we designed an antisense peptide using mRNA transcription of the epitope sequence in the 3'→5' direction and computational screening of potential paratope structures with BLAST; Thirdly, we evaluated sense-antisense (epitope-paratope) peptide binding and affinity by means of fluorescence spectroscopy and microscale thermophoresis. Both methods showed similar Kd values of 850 and 816 µM, respectively. The advantages of the methods were: fast screening with a small quantity of the sample needed, and measurements done within the range of physicochemical parameters resembling physiological conditions. Antisense peptides targeting specific erythropoietin region(s) could be used for the development of new immunochemical methods. Selected antisense peptides with optimal affinity are potential lead compounds for the development of novel diagnostic substances, biopharmaceuticals and vaccines. PMID:24865486

  3. Anaemia in systemic lupus erythematosus: aetiological profile and the role of erythropoietin

    PubMed Central

    Voulgarelis, M.; Kokori, S.; Ioannidis, J.; Tzioufas, A.; Kyriaki, D.; Moutsopoulos, H.

    2000-01-01

    OBJECTIVE—To study the prevalence of different causes of anaemia in patients with systemic lupus erythematosus (SLE) and their associations with immunological and clinical parameters and to evaluate the contribution of erythropoietin (Epo) and anti-erythropoietin (anti-Epo) autoantibodies to the development of SLE anaemia.
METHODS—132 SLE patients with anaemia (defined as haemoglobin of 12 g/dl or less for women and 13.5 g/dl or less for men) from among a total of 345 consecutive SLE patients were prospectively enrolled into the study. Standard haematological and immunological tests were performed and serum Epo and anti-Epo antibodies were assayed.
RESULTS—The identified causes were anaemia of chronic disease (ACD) n=49 (37.1%), iron deficiency anaemia (IDA) n=47 (35.6%), autoimmune haemolytic anaemia (AHA) n=19 (14.4%) and other causes n=17 (12.9%). There was significant heterogeneity in the severity of anaemia between the four groups (p<0.01) with AHA cases being on average more severe. The proportion of patients with anticardiolipin antibodies, low complement levels and anti-dsDNA differed significantly among the four groups; these markers were particularly common in patients with AHA, and uncommon in patients with IDA. Twenty one of 100 tested patients had anti-Epo antibodies. Such antibodies were seen practically only in patients with ACD (odds ratio 3.1, p=0.041) and in patients with high lupus activity (ECLAM) scores (odds ratio 1.27 per point, p=0.055). Epo response was inadequate in 42.4% and 41.2% of patients with ACD and AHA, respectively.
CONCLUSIONS—Anaemia in SLE usually takes the form of ACD and IDA, however autoimmune haemolysis is not uncommon. SLE patients with different causes of anaemia differ in regard to several immunological parameters. Epo response is blunted in anaemic SLE patients, particularly those with ACD and AHA.

 PMID:10700431

  4. Hypoxic pulmonary vasoconstriction, carotid body function and erythropoietin production in adult rats perinatally exposed to hyperoxia

    PubMed Central

    Prieto-Lloret, Jesus; Ramirez, Maria; Olea, Elena; Moral-Sanz, Javier; Cogolludo, Angel; Castañeda, Javier; Yubero, Sara; Agapito, Teresa; Gomez-Niño, Angela; Rocher, Asuncion; Rigual, Ricardo; Obeso, Ana; Perez-Vizcaino, Francisco; González, Constancio

    2015-01-01

    Adult mammalians possess three cell systems that are activated by acute bodily hypoxia: pulmonary artery smooth muscle cells (PASMC), carotid body chemoreceptor cells (CBCC) and erythropoietin (EPO)-producing cells. In rats, chronic perinatal hyperoxia causes permanent carotid body (CB) atrophy and functional alterations of surviving CBCC. There are no studies on PASMC or EPO-producing cells. Our aim is to define possible long-lasting functional changes in PASMC or EPO-producing cells (measured as EPO plasma levels) and, further, to analyse CBCC functional alterations. We used 3- to 4-month-old rats born and reared in a normal atmosphere or exposed to perinatal hyperoxia (55–60% O2 for the last 5–6 days of pregnancy and 4 weeks after birth). Perinatal hyperoxia causes an almost complete loss of hypoxic pulmonary vasoconstriction (HPV), which was correlated with lung oxidative status in early postnatal life and prevented by antioxidant supplementation in the diet. O2-sensitivity of K+ currents in the PASMC of hyperoxic animals is normal, indicating that their inhibition is not sufficient to trigger HPV. Perinatal hyperoxia also abrogated responses elicited by hypoxia on catecholamine and cAMP metabolism in the CB. An increase in EPO plasma levels elicited by hypoxia was identical in hyperoxic and control animals, implying a normal functioning of EPO-producing cells. The loss of HPV observed in adult rats and caused by perinatal hyperoxia, comparable to oxygen therapy in premature infants, might represent a previously unrecognized complication of such a medical intervention capable of aggravating medical conditions such as regional pneumonias, atelectases or general anaesthesia in adult life. Key points Adult animals that have been perinatally exposed to oxygen-rich atmospheres (hyperoxia), recalling those used for oxygen therapy in infants, exhibit a loss of hypoxic pulmonary vasoconstriction, whereas vasoconstriction elicited by depolarizing agents is

  5. Erythropoietin improves the accumulation and therapeutic effects of carboplatin by enhancing tumor vascularization and perfusion.

    PubMed

    Doleschel, Dennis; Rix, Anne; Arns, Susanne; Palmowski, Karin; Gremse, Felix; Merkle, Ruth; Salopiata, Florian; Klingmüller, Ursula; Jarsch, Michael; Kiessling, Fabian; Lederle, Wiltrud

    2015-01-01

    Recombinant human erythropoietin (rhuEpo) is currently under debate for the treatment of chemotherapy-induced anemia due to clinical trials showing adverse effects in Epo-treated patients and the discovery of the erythropoietin-receptor (EpoR) in tumor and endothelial cells. Here, using Epo-Cy5.5 as theranostic near-infrared fluorescent probe we analyzed the effects of rhuEpo as co-medication to carboplatin in non-small-cell-lung-cancer (NSCLC)-xenografts with different tumor cell EpoR-expression (H838 ~8-fold higher than A549). Nude mice bearing subcutaneous A549 and H838 NSCLC-xenografts received either only carboplatin or carboplatin and co-medication of rhuEpo in two different doses. Tumor sizes and relative blood volumes (rBV) were longitudinally measured by 3D-contrast-enhanced ultrasound (3D-US). Tumoral EpoR-levels were determined by combined fluorescence molecular tomography (FMT)/ micro computed tomography (µCT) hybrid imaging. We found that rhuEpo predominantly acted on the tumor endothelium. In both xenografts, rhuEpo co-medication significantly increased vessel densities, diameters and the amount of perfused vessels. Accordingly, rhuEpo induced EpoR-phoshorylation and stimulated proliferation of endothelial cells. However, compared with solely carboplatin-treated tumors, tumor growth was significantly slower in the groups co-medicated with rhuEpo. This is explained by the Epo-mediated vascular remodeling leading to improved drug delivery as obvious by a more than 2-fold higher carboplatin accumulation and significantly enhanced tumor apoptosis. In addition, co-medication of rhuEpo reduced tumor hypoxia and diminished intratumoral EpoR-levels which continuously increased during carboplatin (Cp) -treatment. These findings suggest that co-medication of rhuEpo in well balanced doses can be used to improve the accumulation of anticancer drugs. Doses and indications may be personalized and refined using theranostic EpoR-probes.

  6. Nitric oxide and hypoxia stimulate erythropoietin receptor via MAPK kinase in endothelial cells.

    PubMed

    Cokic, Bojana B Beleslin; Cokic, Vladan P; Suresh, Sukanya; Wirt, Stacey; Noguchi, Constance Tom

    2014-03-01

    Erythropoietin receptor (EPOR) expression level determines the extent of erythropoietin (EPO) response. Previously we showed that EPOR expression in endothelial cells is increased at low oxygen tension and that EPO stimulation of endothelial cells during hypoxia can increase endothelial nitric oxide (NO) synthase (eNOS) expression and activation as well as NO production. We now observe that while EPO can stimulate NO production, NO in turn can regulate EPOR expression. Human umbilical vein endothelial cells (HUVEC) treated with 10-50 μM of NO donor diethylenetriamine NONOate (DETANO) for 24h showed significant induction of EPOR gene expression at 5% and 2% of oxygen. Also human bone marrow microvascular endothelial cell line (TrHBMEC) cultured at 21 and 2% oxygen with 50 μM DETANO demonstrated a time and oxygen dependent induction of EPOR mRNA expression after 24 and 48 h, particularly at low oxygen tension. EPOR protein was also induced by DETANO at 2% oxygen in TrHBMEC and HUVEC. The activation of signaling pathways by NO donor stimulation appeared to be distinct from EPO stimulation. In reporter gene assays, DETANO treatment of HeLa cells at 2% oxygen increased EPOR promoter activity indicated by a 48% increase in luciferase activity with a 2 kb EPOR promoter fragment and a 71% increase in activity with a minimal EPOR promoter fragment containing 0.2 kb 5'. We found that DETANO activated MAPK kinase in TrHBMEC both in normoxia and hypoxia, while MAPK kinase inhibition showed significant reduction of EPOR mRNA gene expression at low oxygen tension, suggesting MAPK involvement in NO mediated induction of EPOR. Furthermore, DETANO stimulated Akt anti-apoptotic activity after 30 min in normoxia, whereas it inhibited Akt phosphorylation in hypoxia. In contrast, EPO did not significantly increase MAPK activity while EPO stimulated Akt phosphorylation in TrHBMEC in normoxia and hypoxia. These observations provide a new effect of NO on EPOR expression to enhance EPO

  7. Erythropoietin Improves the Accumulation and Therapeutic Effects of Carboplatin by Enhancing Tumor Vascularization and Perfusion

    PubMed Central

    Doleschel, Dennis; Rix, Anne; Arns, Susanne; Palmowski, Karin; Gremse, Felix; Merkle, Ruth; Salopiata, Florian; Klingmüller, Ursula; Jarsch, Michael; Kiessling, Fabian; Lederle, Wiltrud

    2015-01-01

    Recombinant human erythropoietin (rhuEpo) is currently under debate for the treatment of chemotherapy-induced anemia due to clinical trials showing adverse effects in Epo-treated patients and the discovery of the erythropoietin-receptor (EpoR) in tumor and endothelial cells. Here, using Epo-Cy5.5 as theranostic near-infrared fluorescent probe we analyzed the effects of rhuEpo as co-medication to carboplatin in non-small-cell-lung-cancer (NSCLC)-xenografts with different tumor cell EpoR-expression (H838 ~8-fold higher than A549). Nude mice bearing subcutaneous A549 and H838 NSCLC-xenografts received either only carboplatin or carboplatin and co-medication of rhuEpo in two different doses. Tumor sizes and relative blood volumes (rBV) were longitudinally measured by 3D-contrast-enhanced ultrasound (3D-US). Tumoral EpoR-levels were determined by combined fluorescence molecular tomography (FMT)/ micro computed tomography (µCT) hybrid imaging. We found that rhuEpo predominantly acted on the tumor endothelium. In both xenografts, rhuEpo co-medication significantly increased vessel densities, diameters and the amount of perfused vessels. Accordingly, rhuEpo induced EpoR-phoshorylation and stimulated proliferation of endothelial cells. However, compared with solely carboplatin-treated tumors, tumor growth was significantly slower in the groups co-medicated with rhuEpo. This is explained by the Epo-mediated vascular remodeling leading to improved drug delivery as obvious by a more than 2-fold higher carboplatin accumulation and significantly enhanced tumor apoptosis. In addition, co-medication of rhuEpo reduced tumor hypoxia and diminished intratumoral EpoR-levels which continuously increased during carboplatin (Cp) -treatment. These findings suggest that co-medication of rhuEpo in well balanced doses can be used to improve the accumulation of anticancer drugs. Doses and indications may be personalized and refined using theranostic EpoR-probes. PMID:26000061

  8. Prognostic Impact of Erythropoietin Expression and Erythropoietin Receptor Expression on Locoregional Control and Survival of Patients Irradiated for Stage II/III Non-Small-Cell Lung Cancer

    SciTech Connect

    Rades, Dirk; Setter, Cornelia; Dahl, Olav; Schild, Steven E.; Noack, Frank

    2011-06-01

    Purpose: Prognostic factors can guide the physician in selecting the optimal treatment for an individual patient. This study investigates the prognostic value of erythropoietin (EPO) and EPO receptor (EPO-R) expression of tumor cells for locoregional control and survival in non-small-cell lung cancer (NSCLC) patients. Methods and Materials: Fourteen factors were investigated in 62 patients irradiated for stage II/III NSCLC, as follows: age, gender, Karnofsky performance score (KPS), histology, grading, TNM/American Joint Committee on Cancer (AJCC) stage, surgery, chemotherapy, pack years (average number of packages of cigarettes smoked per day multiplied by the number of years smoked), smoking during radiotherapy, hemoglobin levels during radiotherapy, EPO expression, and EPO-R expression. Additionally, patients with tumors expressing both EPO and EPO-R were compared to those expressing either EPO or EPO-R and to those expressing neither EPO nor EPO-R. Results: On univariate analysis, improved locoregional control was associated with AJCC stage II cancer (p < 0.048), surgery (p < 0.042), no smoking during radiotherapy (p = 0.024), and no EPO expression (p = 0.001). A trend was observed for a KPS of >70 (p = 0.08), an N stage of 0 to 1 (p = 0.07), and no EPO-R expression (p = 0.10). On multivariate analysis, AJCC stage II and no EPO expression remained significant. No smoking during radiotherapy was almost significant. On univariate analysis, improved survival was associated with N stage 0 to 1 (p = 0.009), surgery (p = 0.039), hemoglobin levels of {>=}12 g/d (p = 0.016), and no EPO expression (p = 0.001). On multivariate analysis, N stage 0 to 1 and no EPO expression maintained significance. Hemoglobin levels of {>=}12 g/d were almost significant. On subgroup analyses, patients with tumors expressing both EPO and EPO-R had worse outcomes than those expressing either EPO or EPO-R and those expressing neither EPO nor RPO-R. Conclusions: EPO expression of tumor cells

  9. A "classical" homodimeric erythropoietin receptor is essential for the antiapoptotic effects of erythropoietin on differentiated neuroblastoma SH-SY5Y and pheochromocytoma PC-12 cells.

    PubMed

    Um, Moonkyoung; Gross, Alec W; Lodish, Harvey F

    2007-03-01

    The hematopoietic cytokine erythropoietin (Epo) exerts cytoprotective effects on several types of neuronal cells both in vivo and in culture. Detailed molecular mechanisms underlying this phenomenon have not been elucidated and even the identity of the cytoprotective Epo receptors in neuronal cells is controversial. Here we show that Epo prevents staurosporine-induced apoptosis of differentiated human neuroblastoma SH-SY5Y cells, and activates the STAT5, AKT and MAPK signaling pathways. Differentiated SH-SY5Y cells have fewer than 50 high affinity Epo surface binding sites per cell, which could not be detected by standard assays measuring binding of 125I-labeled Epo. However, by measuring endocytosis of 125I-Epo, we could reliably quantify very small numbers of high-affinity Epo surface binding sites. Using SH-SY5Y cells stably expressing an Epo receptor (EpoR) shRNA and thus lacking detectable EpoR expression, we show that high affinity binding of Epo to these neuronal cells is mediated by the hematopoietic EpoR, and that this EpoR is also essential for the antiapoptotic activity of Epo. In contrast, a mutant Epo that has an intact binding site 1 but a non-functional binding site 2 and hence binds only to one cell surface EpoR molecule ("site 2" Epo mutant) displays significantly lower antiapoptotic activity than wild-type Epo. Furthermore, expression of the GM-CSF/IL-3/IL-5 receptor common beta chain, which was proposed to be responsible for the cytoprotective activity of Epo on certain types of neuronal cells, was undetectable in differentiated SH-SY5Y cells. Epo also alleviated staurosporine-induced apoptosis of rat PC-12 pheochromocytoma cells while the R103A "site 2" Epo mutant did not, and we could not detect expression of the common beta chain in PC-12 cells. Together our results indicate that Epo exerts its antiapoptotic effects on differentiated SH-SY5Y and PC-12 cells through the standard stoichiometry of one molecule of Epo binding to two EpoR subunits

  10. HIF‑1 signaling pathway involving iNOS, COX‑2 and caspase‑9 mediates the neuroprotection provided by erythropoietin in the retina of chronic ocular hypertension rats.

    PubMed

    Gui, Dongmei; Li, Yanfeng; Chen, Xiaolong; Gao, Dianwen; Yang, Yang; Li, Xun

    2015-02-01

    This study aimed to investigate the impacts of erythropoietin (EPO) on the electroretinogram b‑wave (ERG‑b), and on the mRNA and protein expression levels of hypoxia‑inducible factor‑1α (HIF‑1α), inducible nitric oxide synthase (iNOS), cyclooxygenase‑2 (COX‑2) and caspase‑9 in chronic ocular hypertension rats. Episcleral vein cauterization (EVC) was used to establish the chronic ocular hypertension rat model based on the intraocular pressure (IOP) value. ERG‑b and mRNA and protein expression levels of HIF‑1α, iNOS, COX‑2 and caspase‑9 in normal, EVC‑treated and EVC combined with EPO (EVC+EPO)‑treated rats were measured by electroretinography, RT‑PCR and western blotting, respectively. Moreover, the correlations of HIF‑1α with IOP, ERG‑b, iNOS, COX‑2 and caspase‑9 were evaluated. The mRNA and protein expression levels of HIF‑1α, iNOS, COX‑2 and caspase‑9 in EVC‑treated rats were increased significantly compared with normal rats. The peak expression levels of HIF‑1α, iNOS, COX‑2 and caspase‑9 were respectively obtained 7, 7, 7 and 14 days postoperatively. Compared with EVC‑treated rats, EPO administration weakened the mRNA and protein expression levels of HIF‑1α, iNOS, COX‑2 and caspase‑9. The mRNA expression level of HIF‑1α demonstrated a significant positive correlation with IOP and ERG‑b. HIF‑1α was positively correlated with iNOS, COX‑2 and caspase‑9 at the mRNA and protein levels. The protective effect of EPO on the retina of chronic ocular hypertension rats may be mediated by the HIF‑1 signaling pathway involving iNOS, COX‑2 and caspase‑9.

  11. HIF‑1 signaling pathway involving iNOS, COX‑2 and caspase‑9 mediates the neuroprotection provided by erythropoietin in the retina of chronic ocular hypertension rats.

    PubMed

    Gui, Dongmei; Li, Yanfeng; Chen, Xiaolong; Gao, Dianwen; Yang, Yang; Li, Xun

    2015-02-01

    This study aimed to investigate the impacts of erythropoietin (EPO) on the electroretinogram b‑wave (ERG‑b), and on the mRNA and protein expression levels of hypoxia‑inducible factor‑1α (HIF‑1α), inducible nitric oxide synthase (iNOS), cyclooxygenase‑2 (COX‑2) and caspase‑9 in chronic ocular hypertension rats. Episcleral vein cauterization (EVC) was used to establish the chronic ocular hypertension rat model based on the intraocular pressure (IOP) value. ERG‑b and mRNA and protein expression levels of HIF‑1α, iNOS, COX‑2 and caspase‑9 in normal, EVC‑treated and EVC combined with EPO (EVC+EPO)‑treated rats were measured by electroretinography, RT‑PCR and western blotting, respectively. Moreover, the correlations of HIF‑1α with IOP, ERG‑b, iNOS, COX‑2 and caspase‑9 were evaluated. The mRNA and protein expression levels of HIF‑1α, iNOS, COX‑2 and caspase‑9 in EVC‑treated rats were increased significantly compared with normal rats. The peak expression levels of HIF‑1α, iNOS, COX‑2 and caspase‑9 were respectively obtained 7, 7, 7 and 14 days postoperatively. Compared with EVC‑treated rats, EPO administration weakened the mRNA and protein expression levels of HIF‑1α, iNOS, COX‑2 and caspase‑9. The mRNA expression level of HIF‑1α demonstrated a significant positive correlation with IOP and ERG‑b. HIF‑1α was positively correlated with iNOS, COX‑2 and caspase‑9 at the mRNA and protein levels. The protective effect of EPO on the retina of chronic ocular hypertension rats may be mediated by the HIF‑1 signaling pathway involving iNOS, COX‑2 and caspase‑9. PMID:25370745

  12. Targeted delivery of erythropoietin by transcranial focused ultrasound for neuroprotection against ischemia/reperfusion-induced neuronal injury: a long-term and short-term study.

    PubMed

    Wu, Sheng-Kai; Yang, Ming-Tao; Kang, Kai-Hsiang; Liou, Houng-Chi; Lu, Dai-Hua; Fu, Wen-Mei; Lin, Win-Li

    2014-01-01

    Erythropoietin (EPO) is a neuroprotective agent against cerebral ischemia/reperfusion (I/R)-induced brain injury. However, its crossing of blood-brain barrier is limited. Focused ultrasound (FUS) sonication with microbubbles (MBs) can effectively open blood-brain barrier to boost the vascular permeability. In this study, we investigated the effects of MBs/FUS on extending the therapeutic time window of EPO and its neuroprotective effects in both acute and chronic phases. Male Wistar rats were firstly subjected to two common carotid arteries and right middle cerebral artery occlusion (three vessels occlusion, 3VO) for 50 min, and then the rats were treated with hEPO (human recombinant EPO, 5000 IU/kg) with or without MBs/FUS at 5 h after occlusion/reperfusion. Acute phase investigation (I/R, I/R+MBs/FUS, I/R+hEPO, and I/R+hEPO+MBs/FUS) was performed 24 h after I/R; chronic tests including cylinder test and gait analysis were performed one month after I/R. The experimental results showed that MBs/FUS significantly increased the cerebral content of EPO by bettering vascular permeability. In acute phase, both significant improvement of neurological score and reduction of infarct volume were found in the I/R+hEPO+MBs/FUS group, as compared with I/R and I/R+hEPO groups. In chronic phase, long-term behavioral recovery and neuronal loss in brain cortex after I/R injury was significantly improved in the I/R+hEPO+MBs/FUS group. This study indicates that hEPO administration with MBs/FUS sonication even at 5 h after occlusion/reperfusion can produce a significant neuroprotection. PMID:24587228

  13. Erythropoietin attenuates renal and pulmonary injury in polymicrobial induced-sepsis through EPO-R, VEGF and VEGF-R2 modulation.

    PubMed

    Heitrich, Mauro; García, Daiana Maria de Los Ángeles; Stoyanoff, Tania Romina; Rodríguez, Juan Pablo; Todaro, Juan Santiago; Aguirre, María Victoria

    2016-08-01

    Sepsis remains the most important cause of acute kidney injury (AKI) and acute lung injury (ALI) in critically ill patients. The cecal ligation and puncture (CLP) model in experimental mice reproduces most of the clinical features of sepsis. Erythropoietin (EPO) is a well-known cytoprotective multifunctional hormone, which exerts anti-inflammatory, anti-oxidant, anti-apoptotic and pro-angiogenic effects in several tissues. The aim of this study was to evaluate the underlying mechanisms of EPO protection through the expression of the EPO/EPO receptor (EPO-R) and VEGF/VEF-R2 systems in kidneys and lungs of mice undergoing CLP-induced sepsis. Male inbred Balb/c mice were divided in three experimental groups: Sham, CLP, and CLP+EPO (3000IU/kg sc). Assessment of renal functional parameters, survival, histological examination, immunohistochemistry and/or Western blottings of EPO-R, VEGF and VEGF-R2 were performed at 18h post-surgery. Mice demonstrated AKI by elevation of serum creatinine and renal histologic damage. EPO treatment attenuates renal dysfunction and ameliorates kidney histopathologic changes. Additionally, EPO administration attenuates deleterious septic damage in renal cortex through the overexpression of EPO-R in tubular interstitial cells and the overexpression of the pair VEGF/VEGF-R2. Similarly CLP- induced ALI, as evidenced by parenchymal lung histopathologic alterations, was ameliorated through pulmonary EPO-R, VEGF and VEGF-R2 over expression suggesting and improvement in endothelial survival and functionality. This study demonstrates that EPO exerts protective effects in kidneys and lungs in mice with CLP-induced sepsis through the expression of EPO-R and the regulation of the VEGF/VEGF-R2 pair. PMID:27470403

  14. Restoration of Haemoglobin Level Using Hydrodynamic Gene Therapy with Erythropoietin Does Not Alleviate the Disease Progression in an Anaemic Mouse Model for TGFβ1-Induced Chronic Kidney Disease

    PubMed Central

    Pedersen, Lea; Wogensen, Lise; Marcussen, Niels; Cecchi, Claudia R.; Dalsgaard, Trine; Dagnæs-Hansen, Frederik

    2015-01-01

    Erythropoietin, Epo, is a 30.4 kDa glycoprotein hormone produced primarily by the fetal liver and the adult kidney. Epo exerts its haematopoietic effects by stimulating the proliferation and differentiation of erythrocytes with subsequent improved tissue oxygenation. Epo receptors are furthermore expressed in non-haematopoietic tissue and today, Epo is recognised as a cytokine with many pleiotropic effects. We hypothesize that hydrodynamic gene therapy with Epo can restore haemoglobin levels in anaemic transgenic mice and that this will attenuate the extracellular matrix accumulation in the kidneys. The experiment is conducted by hydrodynamic gene transfer of a plasmid encoding murine Epo in a transgenic mouse model that overexpresses TGF-β1 locally in the kidneys. This model develops anaemia due to chronic kidney disease characterised by thickening of the glomerular basement membrane, deposition of mesangial matrix and mild interstitial fibrosis. A group of age matched wildtype littermates are treated accordingly. After a single hydrodynamic administration of plasmid DNA containing murine EPO gene, sustained high haemoglobin levels are observed in both transgenic and wildtype mice from 7.5 ± 0.6 mmol/L to 9.4 ± 1.2 mmol/L and 10.7 ± 0.3 mmol/L to 15.5 ± 0.5 mmol/L, respectively. We did not observe any effects in the thickness of glomerular or tubular basement membrane, on the expression of different collagen types in the kidneys or in kidney function after prolonged treatment with Epo. Thus, Epo treatment in this model of chronic kidney disease normalises haemoglobin levels but has no effect on kidney fibrosis or function. PMID:26046536

  15. Intravitreal injection of erythropoietin protects against retinal vascular regression at the early stage of diabetic retinopathy in streptozotocin-induced diabetic rats.

    PubMed

    Mitsuhashi, Junko; Morikawa, Shunichi; Shimizu, Kazuhiko; Ezaki, Taichi; Yasuda, Yoshiko; Hori, Sadao

    2013-01-01

    A single intravitreal injection of erythropoietin (EPO) (50 ng/eye) or phosphate-buffered saline was administered to 5-week-old Sprague-Dawley rats at the onset of diabetes mellitus (DM) to determine and evaluate the protective effect of EPO on retinal microvessels. DM was induced by an intraperitoneal injection of streptozotocin (STZ; 60 mg/kg body weight). Morphological changes in microvessels in flat retinal preparations were evaluated during the subsequent 4 weeks by three-dimensional imaging of all blood vessels stained with fluorescein isothiocyanate-conjugated tomato lectin, following immunofluorescence techniques. No marked differences were observed in the shape or density of retinal vessels and the number of retinal capillary branches of the four groups [control, EPO, DM, and DM/EPO] up to 4 weeks after STZ administration. We also observed unique type IV collagen-positive filamentous structures that lacked both cellular elements and blood circulation (lectin-/type IV+ acellular strands), suggesting regressed vessel remnants. The lectin-/type IV+ acellular strands were detected soon after the onset of DM in the diabetic rats, and the number of these structures increased in the DM group (P < 0.01). A single intravitreal injection of EPO caused a significant reduction in the number of lectin-/type IV+ acellular strands to levels observed in the control group. However, the lectin-/type IV+ acellular strands were observed in the central area of the retina near the optic disc in all four groups. Intravitreal injection of EPO resulted in downregulation of the EPO receptor, vascular endothelial growth factor (VEGF), and VEGF receptor at 4 weeks. We conclude that EPO may play a primary role against the progression of diabetic retinopathy by reducing blood vessel degeneration at a very early disease stage. PMID:23178551

  16. Calmodulin physically interacts with the erythropoietin receptor and enhances Jak2-mediated signaling

    SciTech Connect

    Kakihana, Kazuhiko; Yamamoto, Masahide; Iiyama, Mitsuko; Miura, Osamu . E-mail: miura.hema@tmd.ac.jp

    2005-09-23

    Stimulation of the erythropoietin receptor (EpoR) induces a transient increase in intracellular Ca{sup 2+} level as well as activation of the Jak2 tyrosine kinase to stimulate various downstream signaling pathways. Here, we demonstrate that the universal Ca{sup 2+} receptor calmodulin (CaM) binds EpoR in a Ca{sup 2+}-dependent manner in vitro. Binding studies using various EpoR mutants in hematopoietic cells showed that CaM binds the membrane-proximal 65-amino-acid cytoplasmic region (amino acids 258-312) of EpoR that is critical for activation of Jak2-mediated EpoR signaling. Structurally unrelated CaM antagonists, W-13 and CMZ, inhibited activation of Jak2-mediated EpoR signaling pathways, whereas W-12, a W-13 analog, did not show any significant inhibitory effect. Moreover, overexpression of CaM augmented Epo-induced tyrosine phosphorylation of the EpoR. W-13, but not W-12, also inhibited Epo-induced proliferation and survival. Together, these results indicate that CaM binds to the membrane-proximal EpoR cytoplasmic region and plays an essential role in activation of Jak2-mediated EpoR signaling.

  17. Erythropoietin restores C-fiber function and prevents pressure ulcer formation in diabetic mice.

    PubMed

    Demiot, Claire; Sarrazy, Vincent; Javellaud, James; Gourloi, Loriane; Botelle, Laurent; Oudart, Nicole; Achard, Jean-Michel

    2011-11-01

    Pressure-induced vasodilatation (PIV), a cutaneous physiological neurovascular (C-fiber/endothelium) mechanism, is altered in diabetes and could possibly contribute to pressure ulcer development. We wanted to determine whether recombinant human erythropoietin (rhEPO), which has protective neurovascular effects, could prevent PIV alteration and pressure ulcer formation. We developed a skin pressure ulcer model in mice by applying two magnetic plates to the dorsal skin. This induced significant stage 2 ulcers (assessed visually and histologically) in streptozotocin-treated mice with 8 weeks of diabetes compared with very few in controls. Control and streptozotocin mice received either no treatment or systematic rhEPO (3,000 UI kg(-1) intraperitoneally, twice a week) during the last 2 weeks of diabetes. After 8 weeks of diabetes, we assessed ulcer development, PIV, endothelium-dependent vasodilation, C-fiber-mediated nociception threshold, and skin innervation density. Pretreatment with rhEPO fully prevented ulcer development in streptozotocin mice and also fully restored C-fiber nociception, skin innervation density, and significantly improved PIV, but had no effect on endothelium-dependent vasodilation. Our finding that rhEPO treatment protects the skin against pressure-induced ulcers in diabetic mice encourages evaluation of the therapeutic potential for non-hematopoietic analogs of EPO in preventing neuropathic diabetic ulcers.

  18. Distinct subpopulations of FOXD1 stroma-derived cells regulate renal erythropoietin

    PubMed Central

    Liu, Qingdu; Binns, Thomas C.; Davidoff, Olena; Kapitsinou, Pinelopi P.; Pfaff, Andrew S.; Olauson, Hannes; Fogo, Agnes B.; Fong, Guo-Hua; Gross, Kenneth W.

    2016-01-01

    Renal peritubular interstitial fibroblast-like cells are critical for adult erythropoiesis, as they are the main source of erythropoietin (EPO). Hypoxia-inducible factor 2 (HIF-2) controls EPO synthesis in the kidney and liver and is regulated by prolyl-4-hydroxylase domain (PHD) dioxygenases PHD1, PHD2, and PHD3, which function as cellular oxygen sensors. Renal interstitial cells with EPO-producing capacity are poorly characterized, and the role of the PHD/HIF-2 axis in renal EPO-producing cell (REPC) plasticity is unclear. Here we targeted the PHD/HIF-2/EPO axis in FOXD1 stroma-derived renal interstitial cells and examined the role of individual PHDs in REPC pool size regulation and renal EPO output. Renal interstitial cells with EPO-producing capacity were entirely derived from FOXD1-expressing stroma, and Phd2 inactivation alone induced renal Epo in a limited number of renal interstitial cells. EPO induction was submaximal, as hypoxia or pharmacologic PHD inhibition further increased the REPC fraction among Phd2–/– renal interstitial cells. Moreover, Phd1 and Phd3 were differentially expressed in renal interstitium, and heterozygous deficiency for Phd1 and Phd3 increased REPC numbers in Phd2–/– mice. We propose that FOXD1 lineage renal interstitial cells consist of distinct subpopulations that differ in their responsiveness to Phd2 inactivation and thus regulation of HIF-2 activity and EPO production under hypoxia or conditions of pharmacologic or genetic PHD inactivation. PMID:27088801

  19. An alternative to animal testing in the quality control of erythropoietin.

    PubMed

    Zimmermann, H; Gerhard, D; Hothorn, L A; Dingermann, T

    2011-06-01

    A physico-chemical method has been developed as an alternative to the current bioassay in normocythaemic mice for estimating the biological activity of erythropoietin batches. Capillary zone electrophoresis was used for quantification of the isoforms and their substructures were further elucidated by N-glycan mapping techniques. The analytical study was carried out on a total of 40 batches of epoetin beta which were selected to cover an adequate range of precisely established potency values. The relationship between the biological and chemical parameters was evaluated statistically in order to identify suitable covariates for the prediction of the biological activity. Out of several alternatives, a prediction model which is based on the percentages of isoforms per batch and the degree of sialidation was selected and tested. This model is comparable in terms of accuracy to the established in vivo bioassay, but is far superior in terms of precision. Further advantages of the method are improved animal welfare and savings in time and effort. The question whether the prediction model already meets the requirements for replacing the bioassay according to the ICH guideline Q6B is discussed.

  20. Sorting of growth hormone-erythropoietin fusion proteins in rat salivary glands

    SciTech Connect

    Samuni, Yuval Zheng Changyu; Cawley, Niamh X.; Cotrim, Ana P.; Loh, Y. Peng; Baum, Bruce J.

    2008-08-15

    Neuroendocrine and exocrine cells secrete proteins in either a constitutive manner or via the regulated secretory pathway (RSP), but the specific sorting mechanisms involved are not fully understood. After gene transfer to rat salivary glands, the transgenic model proteins human growth hormone (hGH) and erythropoietin (hEpo) are secreted primarily into saliva (RSP; exocrine) and serum (constitutive; endocrine), respectively. We hypothesized that fusion of hGH at either the C-terminus or the N-terminus of hEpo would re-direct hEpo from the bloodstream into saliva. We constructed and expressed two fusion proteins, hEpo-hGH and hGH-hEpo, using serotype 5-adenoviral vectors, and delivered them to rat submandibular glands in vivo via retroductal cannulation. Both the hEpo-hGH and hGH-hEpo fusion proteins, but not hEpo alone, were secreted primarily into saliva (p < 0.0001 and p = 0.0083, respectively). These in vivo studies demonstrate for the first time that hGH, in an N- as well as C-terminal position, influences the secretion of a constitutive pathway protein.

  1. Resistance to Recombinant Human Erythropoietin Therapy in a Rat Model of Chronic Kidney Disease Associated Anemia.

    PubMed

    Garrido, Patrícia; Ribeiro, Sandra; Fernandes, João; Vala, Helena; Rocha-Pereira, Petronila; Bronze-da-Rocha, Elsa; Belo, Luís; Costa, Elísio; Santos-Silva, Alice; Reis, Flávio

    2015-12-25

    This study aimed to elucidate the mechanisms explaining the persistence of anemia and resistance to recombinant human erythropoietin (rHuEPO) therapy in a rat model of chronic kidney disease (CKD)-associated anemia with formation of anti-rHuEPO antibodies. The remnant kidney rat model of CKD induced by 5/6 nephrectomy was used to test a long-term (nine weeks) high dose of rHuEPO (200 UI/kg bw/week) treatment. Hematological and biochemical parameters were evaluated as well as serum and tissue (kidney, liver and/or duodenum) protein and/or gene expression of mediators of erythropoiesis, iron metabolism and tissue hypoxia, inflammation, and fibrosis. Long-term treatment with a high rHuEPO dose is associated with development of resistance to therapy as a result of antibodies formation. In this condition, serum EPO levels are not deficient and iron availability is recovered by increased duodenal absorption. However, erythropoiesis is not stimulated, and the resistance to endogenous EPO effect and to rHuEPO therapy results from the development of a hypoxic, inflammatory and fibrotic milieu in the kidney tissue. This study provides new insights that could be important to ameliorate the current therapeutic strategies used to treat patients with CKD-associated anemia, in particular those that become resistant to rHuEPO therapy.

  2. Tissue-specific regulation of erythropoietin production in the murine kidney, brain, and uterus.

    PubMed

    Chikuma, M; Masuda, S; Kobayashi, T; Nagao, M; Sasaki, R

    2000-12-01

    Erythropoietin (Epo) produced by the kidney regulates erythropoiesis. Recent evidence suggests that Epo in the cerebrum prevents neuron death and Epo in the uterus induces estrogen (E(2))-dependent uterine angiogenesis. To elucidate how Epo expression is regulated in these tissues, ovariectomized mice were given E(2) and/or exposed to hypoxia, and the temporal patterns of Epo mRNA levels were examined. Epo mRNA levels in the kidney and cerebrum were elevated markedly within 4 h after exposure to hypoxia. Although the elevated level of Epo mRNA in the kidney decreased markedly within 8 h despite continuous hypoxia, the high level in the cerebrum was sustained for > or = 24 h, indicating that downregulation operates in the kidney but not in the brain. E(2) transiently induced Epo mRNA in the uterus but not in the kidney and cerebrum. Interestingly, the uterine Epo mRNA was hypoxia inducible only in the presence of E(2). Thus Epo expression appears to be regulated in a tissue-specific manner, endorsing the tissue-specific functions of Epo.

  3. Induction of erythropoietin responsiveness in vitro by a distinct population of bone marrow cells.

    PubMed

    Wagemaker, G; Peters, M F; Bol, S J

    1979-09-01

    Bone marrow contains a small population of primitive erythroid progenitor cells which can be detected by their capacity to form large numbers of erythroid progeny in viscous cultures containing erythropoietin (EP). These cells have been termed erythroid 'burst-forming units' (BFUe). The present study demonstrates that expression of the erythroid differentiation potential of BFUe requires the presence of an activity additional to EP. This activity has been designated as BFA (burst feeder activity). It is shown that the number of BFUe detected and their apparent sensitivity to EP are directly related to the BFA concentration of the cultures. BFA was found to be associated with a population of bone marrow cells of high buoyant density and small volume, which are sensitive to irradiation. The radiation dose-effect curve provided strong evidence that bone marrow BFA is independent of cell proliferation; this was supported by showing that BFA is unaffected by in vivo treatment with hydroxyurea. The findings are compatible with a two-step regulation model for erythroid differentiation in which BFA-induced progeny of BFUe acquire sensitivity to EP. PMID:316359

  4. Low levels of serum erythropoietin in children with endemic hemolytic uremic syndrome.

    PubMed

    Exeni, R; Donato, H; Rendo, P; Antonuccio, M; Rapetti, M C; Grimoldi, I; Exeni, A; de Galvagni, A; Trepacka, E; Amore, A

    1998-04-01

    Serum erythropoietin (EPO) levels were measured in ten previously non-transfused children with hemolytic uremic syndrome (HUS). Complete blood cell count, serum EPO, and renal function tests were carried out upon admission and weekly thereafter. Blood samples were obtained: (1) prior to the first transfusion; (2) after the first transfusion but before recovery from renal failure; (3) during the recovery stage. All patients required transfusions (mean 1.8+/-0.8 per child). Absolute values of EPO correlated positively with the hematocrit during the three stages (r = 0.53, 0.36, and 0.12, respectively) which is opposite to expected results. The observed EPO logarithm/predicted EPO logarithm upon admission was low (0.70+/-0.08), falling further during stage 2 (0.57+/-0.03), but increasing thereafter (0.78+/-0.07) without reaching normal values. The reticulocyte production rate followed a parallel course (0.74+/-0.14, 0.54+/-0.11, and 0.60+/-0.10, respectively). On comparing the observed serum EPO levels with those expected, 9 of 11 pre-transfusion samples showed low values; in stage 2, all samples were below normal; in the recovery phase most (77.8%) were still low. Our results show an inadequate EPO synthesis in children with HUS, which could play an important pathogenic role, since it aggravates the severity of the existing hemolytic anemia; the secondary inhibitory effect of repeated transfusions exacerbates this inadequate synthesis.

  5. Erythropoietin bioassays using fetal mouse liver cells: validations and technical improvements

    SciTech Connect

    Dunn, C.D.; Gibson, L.

    1983-08-01

    Studies are described which were designed to compare an erythropoietin (Ep) bioassay using the uptake of /sup 125/I-deoxyuridine (/sup 125/-I-UdR) into whole cells cultured in micro-titer plates with an established method where the cells were cultured in 1 ml volumes using the incorporation of /sup 59/Fe into heme as the endpoint. The results demonstrate the validity of the substitution of /sup 125/I-UdR uptake into whole cells as a replacement for /sup 59/Fe incorporation into heme as an assay endpoint and the adaptation of the method to a semi-micro scale with automated cell harvesting. A culture time significantly shorter than 24 h is demonstrated not to be of practical benefit. Two methods are proposed to saturate the growth promoting effects of iron-containing transferrin. The performance of semi-micro, serum-containing bioassay employing untreated serum as the test material is shown to be superior to previous systems for the biological measurement of Ep.

  6. Neuronal erythropoietin overexpression protects mice against age-related hearing loss (presbycusis).

    PubMed

    Naldi, Arianne Monge; Belfrage, Celina; Jain, Neha; Wei, Eric T; Martorell, Belén Canto; Gassmann, Max; Vogel, Johannes

    2015-12-01

    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. PMID:26364734

  7. Erythropoietin-mediated neuroprotection in a pediatric mouse model of chronic hypoxia.

    PubMed

    Chung, Eugene; Kong, Xiangmei; Goldberg, Mark P; Stowe, Ann M; Raman, Lakshmi

    2015-06-15

    Chronic hypoxia (CH), a disease state that accounts for significant morbidity and mortality in pediatrics, occurs in many children during critical periods of hippocampal development and cortical myelination. Hippocampal neurogenesis occurs throughout postnatal life and is important for normal development, thus impairment results in long-term cognitive deficits. Erythropoietin (EPO), a drug commonly known for its role in erythrogenesis, has recently been evaluated in neuroprotection in neonatal injury models and preterm brain injury. However, the effects of EPO therapy on hippocampal neurogenesis and myelination in pediatric CH are unknown. We show that CH decreases hippocampal neurogenesis in a pediatric mouse model. This decrease in early and late progenitors, and actively dividing cells is rescued with EPO treatment. Furthermore, we show that CH during this critical time decreases oligodendrocyte progenitor (OPC) populations in the cortex, leading to defective myelination. However, EPO therapy is only able to rescue the OPC but not the loss of mature myelin. Overall, our findings demonstrate that CH in developing mice has significant effects on hippocampal neurogenesis and OPCs, which can be rescued with EPO treatment. Future studies should confirm the role of this FDA-approved therapy in neuroprotection in at-risk pediatric populations. PMID:25899777

  8. Beneficial Effect of Erythropoietin Short Peptide on Acute Traumatic Brain Injury.

    PubMed

    Wang, Bo; Kang, Mitchell; Marchese, Michelle; Rodriguez, Esther; Lu, Wei; Li, Xintong; Maeda, Yasuhiro; Dowling, Peter

    2016-04-01

    There is currently no effective medical treatment for traumatic brain injury (TBI). Beyond the immediate physical damage caused by the initial impact, additional damage evolves due to the inflammatory response that follows brain injury. Here we show that therapy with JM4, a low molecular weight 19-amino acid nonhematopoietic erythropoietin (EPO) peptidyl fragment, containing amino acids 28-46 derived from the first loop of EPO, markedly reduces acute brain injury. Mice underwent controlled cortical injury and received either whole molecule EPO, JM4, or sham-treatment with phosphate-buffered saline. Animals treated with JM4 peptide exhibited a large decrease in number of dead neural cells and a marked reduction in lesion size at both 3 and 8 days postinjury. Therapy with JM4 also led to improved functional recovery and we observed a treatment window for JM4 peptide that remained open for at least 9 h postinjury. The full-length EPO molecule was divided into a series of 6 contiguous peptide segments; the JM4-containing segment and the adjoining downstream region contained the bulk of the death attenuating effects seen with intact EPO molecule following TBI. These findings indicate that the JM4 molecule substantially blocks cell death and brain injury following acute brain trauma and, as such, presents an excellent opportunity to explore the therapeutic potential of a small-peptide EPO derivative in the medical treatment of TBI. PMID:26715414

  9. Erythropoietin, a novel versatile player regulating energy metabolism beyond the erythroid system.

    PubMed

    Wang, Li; Di, Lijun; Noguchi, Constance Tom

    2014-01-01

    Erythropoietin (EPO), the required cytokine for promoting the proliferation and differentiation of erythroid cells to stimulate erythropoiesis, has been reported to act as a pleiotropic cytokine beyond hematopoietic system. The various activities of EPO are determined by the widespread distribution of its cell surface EPO receptor (EpoR) in multiple tissues including endothelial, neural, myoblasts, adipocytes and other cell types. EPO activity has been linked to angiogenesis, neuroprotection, cardioprotection, stress protection, anti-inflammation and especially the energy metabolism regulation that is recently revealed. The investigations of EPO activity in animals and the expression analysis of EpoR provide more insights on the potential of EPO in regulating energy metabolism and homeostasis. The findings of crosstalk between EPO and some important energy sensors and the regulation of EPO in the cellular respiration and mitochondrial function further provide molecular mechanisms for EPO activity in metabolic activity regulation. In this review, we will summarize the roles of EPO in energy metabolism regulation and the activity of EPO in tissues that are tightly associated with energy metabolism. We will also discuss the effects of EPO in regulating oxidative metabolism and mitochondrial function, the interactions between EPO and important energy regulation factors, and the protective role of EPO from stresses that are related to metabolism, providing a brief overview of previously less appreciated EPO biological function in energy metabolism and homeostasis. PMID:25170305

  10. Safety and angiogenic effects of systemic gene delivery of a modified erythropoietin.

    PubMed

    de Lucas Cerrillo, A M; Bond, W S; Rex, T S

    2015-05-01

    Erythropoietin (EPO) is critical for red blood cell production and is also an effective neuroprotective agent. However, it may contribute to pathological angiogenesis. Here we investigate the angiogenic potential of EPO and a mutant form with attenuated erythropoietic activity, EPO-R76E, on primary human retinal microvascular endothelial cells (HRMECs) and in the adult retina. Assays of death, proliferation and tube formation were performed on HRMECs exposed to EPO, EPO-R76E or media alone. Postnatal day-9 wild-type mice were injected intramuscularly with adeno-associated virus vectors expressing either enhanced green fluorescent protein or EpoR76E. At 3 months, levels of EPO-R76E in the eye were quantified, and the health of the retinal vasculature was assessed by fluorescein angiography and isolectin immunolabeling. Immunohistochemistry, histology and electroretinogram (ERG) assessments were performed as measures of retinal health. Neither EPO nor EPO-R76E induced proliferation or tube formation in HRMECs under the conditions used. EPO-R76E decreased HRMEC death in a dose-dependent manner. Long-term systemic gene delivery of EPO-R76E was safe in terms of retinal vasculature, histology and the ERG in vivo. Our results show that EPO-R76E can block HRMEC death, consistent with its role in erythropoiesis and neuroprotection. In addition, long-term gene delivery of EPO-R76E is safe in the adult retina. PMID:25716531

  11. Serum immunoreactive erythropoietin in high altitude natives with and without excessive erythrocytosis.

    PubMed

    León-Velarde, F; Monge, C C; Vidal, A; Carcagno, M; Criscuolo, M; Bozzini, C E

    1991-05-01

    We report the estimation of blood hemoglobin (Hb), arterial blood oxygen saturation (SaO2), and serum immunoreactive erythropoietin (siEPO) in a group of Peruvian workers residing in Cerro de Pasco at 4300 m showing "excessive erythrocytosis" (EE, Monge's disease, chronic mountain sickness). These estimates were compared with those of humans residing either in Cerro de Pasco and showing "normal erythrocytosis" (NE) or in Lima (sea level, SL) to determine whether Hb and SaO2 are related to siEPO in high altitude (HA) natives with NE or EE. The three parameters showed statistically significant differences between HA and SL groups--the values in SL being lower. Significant differences were also found between NE and EE groups in Hb and SaO2. There was no statistical difference in siEPo between the two groups. The results indicate, therefore, that HA residents who develop EE are not distinguishable from residents who develop NE on the basis of estimates of siEPO. As a result, siEPO and Hb do not show a dose-response relationship in HA residents, and variation in EPO does not explain the striking variation in Hb at high altitudes.

  12. Erythropoietin, protein, and iron supplementation and the prevention of anaemia of prematurity.

    PubMed Central

    Bechensteen, A G; Hågå, P; Halvorsen, S; Whitelaw, A; Liestøl, K; Lindemann, R; Grøgaard, J; Hellebostad, M; Saugstad, O D; Grønn, M

    1993-01-01

    The effectiveness of recombinant human erythropoietin (r-HuEpo) in raising haemoglobin concentrations in very low birthweight infants was examined in a randomised multicentre study. Twenty nine 'healthy' appropriate for gestational age infants with birth weights 900-1400 g entered the study at 3 weeks of age. All infants received breast milk supplemented with 9 g/l human breast milk protein from 3 to 8 weeks of age. Eighteen mg iron was given daily from week 3 and was doubled if serum iron concentration fell below 16.0 mumol/l. Fourteen infants were randomised to receive 100 U/kg r-HuEpo subcutaneously three times a week from week 3 to week 7; 15 infants served as controls. After one week reticulocyte and haemoglobin concentrations were significantly higher in the r-HuEpo treated group and the haemoglobin values remained significantly higher throughout r-HuEpo treatment and at the concentrations observed in full term infants. No adverse effects were associated with the treatment. In stable very low birthweight infants with optimal iron and protein intakes, moderate dose r-HuEpo can produce significant gains in red cell production that may be clinically useful. PMID:8346946

  13. Polycythemia vera. The in vitro response of normal and abnormal stem cell lines to erythropoietin.

    PubMed Central

    Prchal, J F; Adamson, J W; Murphy, S; Steinmann, L; Fialkow, P J

    1978-01-01

    Bone marrow cells from two glucose-6-phosphate dehydrogenase (G-6-PD) heterozygotes with polycythemia vera were cultured to determine whether progenitors which wre not of the polycythemia vera clone were present, and, if present, which cell lines contributed to the increase in erythroid colonies observed in response to added erythropoietin (ESF). To accomplish this, the G-6-PD isoenzyme activity of individual erythroid colonies was determined. All of the erythroid colonies analyzed in cultures without added ESF, contained the G-6-PD isoenzyme type characteristic of the abnormal clone. With higher ESF concentrations in the culture, however, there was an increase in the colonies that were not of the polycythemia vera clone. Analysis of the ratio of the various types of colonies indicated that normal and polycythemia vera cells are capable of responding to ESF in vitro. In selected patients, this technique permits analysis of the ratios of normal to abnormal cells during the course of the disease, in response to therapy and during late complications, such as myelofibrosis or leukemic transformation. PMID:659576

  14. Erythropoietin-driven proliferation of cells with mutations in the tumor suppressor gene TSC2

    PubMed Central

    Ikeda, Yoshihiko; Taveira-DaSilva, Angelo M.; Pacheco-Rodriguez, Gustavo; Steagall, Wendy K.; El-Chemaly, Souheil; Gochuico, Bernadette R.; May, Rose M.; Hathaway, Olanda M.; Li, Shaowei; Wang, Ji-an; Darling, Thomas N.; Stylianou, Mario

    2011-01-01

    Lymphangioleiomyomatosis (LAM) is characterized by cystic lung destruction, resulting from proliferation of smooth-muscle-like cells, which have mutations in the tumor suppressor genes TSC1 or TSC2. Among 277 LAM patients, severe disease was associated with hypoxia and elevated red blood cell indexes that accompanied reduced pulmonary function. Because high red cell indexes could result from hypoxemia-induced erythropoietin (EPO) production, and EPO is a smooth muscle cell mitogen, we investigated effects of EPO in human cells with genetic loss of tuberin function, and we found that EPO increased proliferation of human TSC2−/−, but not of TSC2+/−, cells. A discrete population of cells grown from explanted lungs was characterized by the presence of EPO receptor and loss of heterozygosity for TSC2, consistent with EPO involvement. In LAM cells from lung nodules, EPO was localized to the extracellular matrix, supporting evidence for activation of an EPO-driven signaling pathway. Although the high red cell mass of LAM patients could be related to advanced disease, we propose that EPO, synthesized in response to episodic hypoxia, may increase disease progression by enhancing the proliferation of LAM cells. PMID:21036916

  15. Cloning and expression of human erythropoietin cDNA in Escherichia coli.

    PubMed Central

    Lee-Huang, S

    1984-01-01

    Human erythropoietin (Ep) cDNA has been cloned in Escherichia coli by using pBR322 as a vector. Polyadenylylated RNA was isolated from selected human renal carcinomas with elevated Ep titers. The presence of Ep mRNA was detected by immunoprecipitation of in vitro translation products with monoclonal antibody to human Ep. Double-stranded cDNA was synthesized and inserted into the Pst I site of pBR322 by homopolymeric dG . dC tailing. The cDNA library was initially screened by colony hybridization with 32P-labeled cDNA synthesized from size-fractionated mRNA enriched in Ep message. Positive colonies were further screened immunologically by in situ radioimmunoassay with monoclonal antibody to human Ep. Three positive clones were identified that express the Ep gene sequences as a beta-lactamase fusion protein. These clones contain inserts of approximately 1400, 600, and 200 base pairs. Human renal Ep mRNA, of which the translation products immunoreact with anti-Ep on immunoblots, was hybrid-selected by plasmid DNA from these recombinants. Purified human Ep competes with 35S-labeled hybrid-selected translation products for antibody binding. Images PMID:6371819

  16. Postnatal Erythropoietin Mitigates Impaired Cerebral Cortical Development Following Subplate Loss from Prenatal Hypoxia-Ischemia.

    PubMed

    Jantzie, Lauren L; Corbett, Christopher J; Firl, Daniel J; Robinson, Shenandoah

    2015-09-01

    Preterm birth impacts brain development and leads to chronic deficits including cognitive delay, behavioral problems, and epilepsy. Premature loss of the subplate, a transient subcortical layer that guides development of the cerebral cortex and axonal refinement, has been implicated in these neurological disorders. Subplate neurons influence postnatal upregulation of the potassium chloride co-transporter KCC2 and maturation of γ-amino-butyric acid A receptor (GABAAR) subunits. We hypothesized that prenatal transient systemic hypoxia-ischemia (TSHI) in Sprague-Dawley rats that mimic brain injury from extreme prematurity in humans would cause premature subplate loss and affect cortical layer IV development. Further, we predicted that the neuroprotective agent erythropoietin (EPO) could attenuate the injury. Prenatal TSHI induced subplate neuronal loss via apoptosis. TSHI impaired cortical layer IV postnatal upregulation of KCC2 and GABAAR subunits, and postnatal EPO treatment mitigated the loss (n ≥ 8). To specifically address how subplate loss affects cortical development, we used in vitro mechanical subplate ablation in slice cultures (n ≥ 3) and found EPO treatment attenuates KCC2 loss. Together, these results show that subplate loss contributes to impaired cerebral development, and EPO treatment diminishes the damage. Limitation of premature subplate loss and the resultant impaired cortical development may minimize cerebral deficits suffered by extremely preterm infants.

  17. Negative Relationship between Erythropoietin Dose and Blood Lead Level in Patients Undergoing Maintenance Hemodialysis

    PubMed Central

    Huang, Wen-Hung; Hsu, Ching-Wei; Weng, Cheng-Hao; Lin-Tan, Dan-Tzu; Yen, Tzung-Hai

    2016-01-01

    The adverse effects of increased blood lead levels have been well discussed. Several antioxidant agents have been reported to offer protection from lead toxicity and to reduce blood lead levels (BLL). Given that erythropoietin (EPO) also has antioxidant properties, the aim of this cross-sectional study was to assess the role of EPO and other clinical variables on BLL in hemodialysis (HD) patients. We recruited 931 maintenance hemodialysis (MHD) patients who had undergone HD for at least 6 months and who had ever received blood lead level (BLL) study. Use of erythropoiesis-stimulating agents followed the The National Kidney Foundation Kidney Disease Outcomes Quality Initiative (NKF KDOQI) Clinical Practice Guideline. We estimated demographic, hematological, nutritional, inflammatory, biochemical, and dialysis-related data based on this study. In the group with EPO, 7% had high BLL. In the group without EPO, 22% had high BLL. From the stepwise liner regression, urban areas, hemodialysis duration, and clearance of urea (KT/Vurea) were positively associated with log BLL. In contrast, diabetes (DM), and monthly EPO dose were negatively associated with log BLL. This study showed that EPO dose might be negatively associated with blood lead levels in patients on maintenance hemodialysis. PMID:27680289

  18. BET bromodomain inhibition rescues erythropoietin differentiation of human erythroleukemia cell line UT7

    SciTech Connect

    Goupille, Olivier; Penglong, Tipparat; Lefevre, Carine; Granger, Marine; Kadri, Zahra; Fucharoen, Suthat; Maouche-Chretien, Leila; Leboulch, Philippe; Chretien, Stany

    2012-12-07

    Highlights: Black-Right-Pointing-Pointer UT7 erythroleukemia cells are known to be refractory to differentiate. Black-Right-Pointing-Pointer Brief JQ1 treatment initiates the first steps of erythroid differentiation program. Black-Right-Pointing-Pointer Engaged UT7 cells then maturate in the presence of erythropoietin. Black-Right-Pointing-Pointer Sustained JQ1 treatment inhibits both proliferation and erythroid differentiation. -- Abstract: Malignant transformation is a multistep process requiring oncogenic activation, promoting cellular proliferation, frequently coupled to inhibition of terminal differentiation. Consequently, forcing the reengagement of terminal differentiation of transformed cells coupled or not with an inhibition of their proliferation is a putative therapeutic approach to counteracting tumorigenicity. UT7 is a human leukemic cell line able to grow in the presence of IL3, GM-CSF and Epo. This cell line has been widely used to study Epo-R/Epo signaling pathways but is a poor model for erythroid differentiation. We used the BET bromodomain inhibition drug JQ1 to target gene expression, including that of c-Myc. We have shown that only 2 days of JQ1 treatment was required to transitory inhibit Epo-induced UT7 proliferation and to restore terminal erythroid differentiation. This study highlights the importance of a cellular erythroid cycle break mediated by c-Myc inhibition before initiation of the erythropoiesis program and describes a new model for BET bromodomain inhibitor drug application.

  19. Erythropoietin-directed erythropoiesis depends on serpin inhibition of erythroblast lysosomal cathepsins.

    PubMed

    Dev, Arvind; Byrne, Susan M; Verma, Rakesh; Ashton-Rickardt, Philip G; Wojchowski, Don M

    2013-02-11

    Erythropoietin (EPO) and its cell surface receptor (EPOR) are essential for red blood cell production and exert important cytoprotective effects on select vascular, immune, and cancer cells. To discover novel EPO action modes, we profiled the transcriptome of primary erythroid progenitors. We report Serpina3g/Spi2A as a major new EPO/EPOR target for the survival of erythroid progenitors. In knockout mice, loss of Spi2A worsened anemia caused by hemolysis, radiation, or transplantation. EPO-induced erythropoiesis also was compromised. In particular, maturing erythroblasts required Spi2A for cytoprotection, with iron and reactive oxygen species as cytotoxic agents. Spi2A defects were ameliorated by cathepsin-B/L inhibition, and by genetic co-deletion of lysosomal cathepsin B. Pharmacological inhibition of cathepsin B/L enhanced EPO-induced red cell formation in normal mice. Overall, we define an unexpected EPO action mode via an EPOR-Spi2A serpin-cathepsin axis in maturing erythroblasts, with lysosomal cathepsins as novel therapeutic targets.

  20. The nonhematopoietic effects of erythropoietin in skin regeneration and repair: from basic research to clinical use.

    PubMed

    Sorg, Heiko; Harder, Yves; Krueger, Christian; Reimers, Kerstin; Vogt, Peter M

    2013-05-01

    Erythropoietin (EPO) is the main regulator of red blood cell production but there exists also a variety of nonhematopoietic properties. More recent data show that EPO is also associated with the protection of tissues suffering from ischemia and reperfusion injury as well as with improved regeneration in various organ systems, in particular the skin. This review highlights the mechanisms of EPO in the different stages of wound healing and the reparative processes in the skin emphasizing pathophysiological mechanisms and potential clinical applications. There is clear evidence that EPO effectively influences all wound-healing phases in a dose-dependent manner. This includes inflammation, tissue, and blood vessel formation as well as the remodeling of the wound. The molecular mechanism is predominantly based on an increased expression of the endothelial and inducible nitric oxide (NO) synthase with a consecutive rapid supply of NO as well as an increased content of vascular endothelial growth factor (VEGF) in the wound. The improved understanding of the functions and regulatory mechanisms of EPO in the context of wound-healing problems and ischemia/reperfusion injury, especially during flap surgery, may lead to new considerations of this growth hormone for its regular clinical application in patients.

  1. A gene responsible for prolyl-hydroxylation of moss-produced recombinant human erythropoietin.

    PubMed

    Parsons, Juliana; Altmann, Friedrich; Graf, Manuela; Stadlmann, Johannes; Reski, Ralf; Decker, Eva L

    2013-01-01

    Recombinant production of pharmaceutical proteins is crucial, not only for personalized medicine. While most biopharmaceuticals are currently produced in mammalian cell culture, plant-made pharmaceuticals gain momentum. Post-translational modifications in plants are similar to those in humans, however, existing differences may affect quality, safety and efficacy of the products. A frequent modification in higher eukaryotes is prolyl-4-hydroxylase (P4H)-catalysed prolyl-hydroxylation. P4H sequence recognition sites on target proteins differ between humans and plants leading to non-human posttranslational modifications of recombinant human proteins produced in plants. The resulting hydroxyprolines display the anchor for plant-specific O-glycosylation, which bears immunogenic potential for patients. Here we describe the identification of a plant gene responsible for non-human prolyl-hydroxylation of human erythropoietin (hEPO) recombinantly produced in plant (moss) bioreactors. Targeted ablation of this gene abolished undesired prolyl-hydroxylation of hEPO and thus paves the way for plant-made pharmaceuticals humanized via glyco-engineering in moss bioreactors.

  2. The Role of Protein Excipient in Driving Antibody Responses to Erythropoietin.

    PubMed

    Christie, Merry; Peritt, David; Torres, Raul M; Randolph, Theodore W; Carpenter, John F

    2015-12-01

    Human serum albumin (HSA) is an excipient present in formulations of several recombinant protein products that are approved for clinical use. We investigated the relative contributions of HSA and HSA particles to the generation of antibody responses against recombinant human erythropoietin (rhEPO) and the excipient HSA itself. Protein samples were characterized before injection for quantities of monomeric proteins, soluble protein aggregates, and nano- and micron-sized particles. rhEPO, containing various concentrations of HSA particles, were injected three times a week for 8 weeks into mice. Hematocrits and the production of anti-rhEPO and anti-HSA antibodies were determined at various time points. Levels of antibodies against rhEPO in mice injected with HSA-containing rhEPO were higher than those in mice treated with HSA-free rhEPO. Mice injected with formulations that contained particles of HSA produced strong anti-HSA antibody responses; whereas these responses were greatly reduced when particle-free formulations were administered. In contrast, anti-rhEPO antibody responses were not affected by the presence of particles.

  3. Population pharmacodynamic analysis of erythropoiesis in preterm infants for determining the anemia treatment potential of erythropoietin.

    PubMed

    Saleh, Mohammad I; Nalbant, Demet; Widness, John A; Veng-Pedersen, Peter

    2013-05-01

    A population pharmacokinetics/pharmacodynamic (PK/PD) model was developed to describe changes in erythropoiesis as a function of plasma erythropoietin (EPO) concentration over the first 30 days of life in preterm infants who developed severe anemia requiring red blood cell (RBC) transfusion. Several covariates were tested as possible factors influencing the responsiveness to EPO. Discarded blood samples in 27 ventilated preterm infants born at 24-29 wk of gestation were used to construct plasma EPO, hemoglobin (Hb), and RBC concentration-time profiles. The amount of Hb removed for laboratory testing and that transfused throughout the study period were recorded. A population PK/PD model accounting for the dynamic Hb changes experienced by these infants was simultaneously fitted to plasma EPO, Hb, and RBC concentrations. A covariate analysis suggested that the erythropoietic efficacy of EPO is increased for preterm infants at later gestational ages. The PD analysis showed a sevenfold difference in maximum Hb production rate dependent on gestational age and indicated that preterm infants, when stimulated by EPO, have the capacity to produce additional Hb that may result in a decrease in RBC transfusions. The present model has utility in clinical trial simulations investigating the treatment potential of erythropoietic stimulating agents in the treatment of anemia of prematurity.

  4. Renal epithelium regulates erythropoiesis via HIF-dependent suppression of erythropoietin.

    PubMed

    Farsijani, Navid M; Liu, Qingdu; Kobayashi, Hanako; Davidoff, Olena; Sha, Feng; Fandrey, Joachim; Ikizler, T Alp; O'Connor, Paul M; Haase, Volker H

    2016-04-01

    The adult kidney plays a central role in erythropoiesis and is the main source of erythropoietin (EPO), an oxygen-sensitive glycoprotein that is essential for red blood cell production. Decreases of renal pO2 promote hypoxia-inducible factor 2-mediated (HIF-2-mediated) induction of EPO in peritubular interstitial fibroblast-like cells, which serve as the cellular site of EPO synthesis in the kidney. It is not clear whether HIF signaling in other renal cell types also contributes to the regulation of EPO production. Here, we used a genetic approach in mice to investigate the role of renal epithelial HIF in erythropoiesis. Specifically, we found that HIF activation in the proximal nephron via induced inactivation of the von Hippel-Lindau tumor suppressor, which targets the HIF-α subunit for proteasomal degradation, led to rapid development of hypoproliferative anemia that was associated with a reduction in the number of EPO-producing renal interstitial cells. Moreover, suppression of renal EPO production was associated with increased glucose uptake, enhanced glycolysis, reduced mitochondrial mass, diminished O2 consumption, and elevated renal tissue pO2. Our genetic analysis suggests that tubulointerstitial cellular crosstalk modulates renal EPO production under conditions of epithelial HIF activation in the kidney.

  5. Erythropoietin prevents endothelial dysfunction in GTP-cyclohydrolase I-deficient hph1 mice.

    PubMed

    dʼUscio, Livius V; Santhanam, Anantha V R; Katusic, Zvonimir S

    2014-12-01

    : In this study, we used the mutant hph1 mouse model, which has deficiency in GTP-cyclohydrolase I (GTPCH I) activity, to test the hypothesis that erythropoietin (EPO) protects aortic wall from oxidative stress induced by uncoupling of endothelial nitric oxide synthase (eNOS). Both GTPCH I activity and tetrahydrobiopterin (BH4) levels were reduced in hph1 mice, whereas 7,8-dihydrobiopterin (7,8-BH2) levels were significantly increased. Furthermore, BH4 deficiency caused increased production of superoxide anion and hydrogen peroxide in the aorta thus resulting in impairment of endothelium-dependent relaxations to acetylcholine. Treatment of hph1 mice with recombinant human EPO (1000 U/kg, subcutaneously for 3 days) significantly decreased superoxide anion production by eNOS and improved BH4 to 7,8-BH2 ratio in aortas. EPO also significantly decreased production of hydrogen peroxide and improved endothelium-dependent relaxations in aortas of hph1 mice. In addition, EPO treatment increased protein expressions of copper-/zinc-superoxide dismutase, manganese-superoxide dismutase, and catalase in the aorta of hph1 mice. Our findings demonstrate that treatment with EPO prevented oxidative stress and endothelial dysfunction caused by eNOS uncoupling. Increased vascular expressions of antioxidants seem to be an important molecular mechanism underlying vascular protection by EPO during chronic BH4 deficiency.

  6. Inhibition of erythropoietin signalling destroys xenografts of ovarian and uterine cancers in nude mice

    PubMed Central

    Yasuda, Y; Musha, T; Tanaka, H; Fujita, Y; Fujita, H; Utsumi, H; Matsuo, T; Masuda, S; Nagao, M; Sasaki, R; Nakamura, Y

    2001-01-01

    We have recently shown that malignant tumours from the ovary and uterus expressed erythropoietin (Epo) and its receptor (EpoR), and that deprivation of Epo signal in tumour blocks induced death of malignant cells and capillary endothelial cells in vitro (Yasuda et al, submitted). These in vitro results prompted us to examine the effect of Epo-signal withdrawal on tumours in vivo. RT-PCR analysis demonstrated the expression of mRNAs for Epo and EpoR in the transplants of uterine and ovarian tumours in nude mice. Then we injected locally anti-Epo antibody or soluble form of EpoR into the transplants. At 12 h, 1, 7 or 14 days after the injection, all transplants were resected and examined macro- and microscopically. Tumour size was reduced in Epo signal-deprived transplants. Immunohistochemical examinations revealed destruction of Epo-responding malignant and capillary endothelial cells through apoptotic death. The degree of tumour regression correlated well with the dose and frequency of the injections. Control xenografts with saline injection or needle insertion showed well-developed tumour masses. This Epo response pathway will have profound implications for our understanding of the development and progression of malignant tumours and for the use of Epo-signal deprivation as an effective therapy. © 2001 Cancer Research Campaign http://www.bjcancer.com PMID:11259101

  7. Packing Density of the Erythropoietin Receptor Transmembrane Domain Correlates with Amplification of Biological Responses

    SciTech Connect

    Becker, Verena; Sengupta, D; Ketteler, Robin; Ullmann, G. Matthias; Smith, Jeremy C; Klingmuller, Ursula

    2008-10-01

    The formation of signal-promoting dimeric or oligomeric receptor complexes at the cell surface is modulated by self-interaction of their transmembrane (TM) domains. To address the importance of TM domain packing density for receptor functionality, we examined a set of asparagine mutants in the TM domain of the erythropoietin receptor (EpoR). We identified EpoR-T242N as a receptor variant that is present at the cell surface similar to wild-type EpoR but lacks visible localization in vesicle-like structures and is impaired in efficient activation of specific signaling cascades. Analysis by a molecular modeling approach indicated an increased interhelical distance for the EpoR-T242N TM dimer. By employing the model, we designed additional mutants with increased or decreased packing volume and confirmed a correlation between packing volume and biological responsiveness. These results propose that the packing density of the TM domain provides a novel layer for fine-tuned regulation of signal transduction and cellular decisions.

  8. Erythropoietin protects myocardin-expressing cardiac stem cells against cytotoxicity of tumor necrosis factor-{alpha}

    SciTech Connect

    Madonna, Rosalinda; Shelat, Harnath; Xue, Qun; Willerson, James T.; De Caterina, Raffaele; Geng, Yong-Jian

    2009-10-15

    Cardiac stem cells are vulnerable to inflammation caused by infarction or ischemic injury. The growth factor, erythropoietin (Epo), ameliorates the inflammatory response of the myocardium to ischemic injury. This study was designed to assess the role of Epo in regulation of expression and activation of the cell death-associated intracellular signaling components in cardiac myoblasts stimulated with the proinflammatory cytokine tumor necrosis factor (TNF)-{alpha}. Cardiac myoblasts isolated from canine embryonic hearts characterized by expression of myocardin A, a promyogenic transcription factor for cardiovascular muscle development were pretreated with Epo and then exposed to TNF-{alpha}. Compared to untreated cells, the Epo-treated cardiac myoblasts exhibited better morphology and viability. Immunoblotting revealed lower levels of active caspase-3 and reductions in iNOS expression and NO production in Epo-treated cells. Furthermore, Epo pretreatment reduced nuclear translocation of NF-{kappa}B and inhibited phosphorylation of inhibitor of kappa B (I{kappa}B) in TNF-{alpha}-stimulated cardiac myoblasts. Thus, Epo protects cardiac myocyte progenitors or myoblasts against the cytotoxic effects of TNF-{alpha} by inhibiting NF-{kappa}B-mediated iNOS expression and NO production and by preventing caspase-3 activation.

  9. Carbamylated erythropoietin protects the kidneys from ischemia-reperfusion injury without stimulating erythropoiesis

    SciTech Connect

    Imamura, Ryoichi; Isaka, Yoshitaka . E-mail: isaka@att.med.osaka-u.ac.jp; Ichimaru, Naotsugu; Takahara, Shiro; Okuyama, Akihiko

    2007-02-16

    Several studies have shown that erythropoietin (EPO) can protect the kidneys from ischemia-reperfusion injury and can raise the hemoglobin (Hb) concentration. Recently, the EPO molecule modified by carbamylation (CEPO) has been identified and was demonstrated to be able to protect several organs without increasing the Hb concentration. We hypothesized that treatment with CEPO would protect the kidneys from tubular apoptosis and inhibit subsequent tubulointerstitial injury without erythropoiesis. The therapeutic effect of CEPO was evaluated using a rat ischemia-reperfusion injury model. Saline-treated kidneys exhibited increased tubular apoptosis with interstitial expression of {alpha}-smooth muscle actin ({alpha}-SMA), while EPO treatment inhibited tubular apoptosis and {alpha}-SMA expression to some extent. On the other hand, CEPO-treated kidneys showed minimal tubular apoptosis with limited expression of {alpha}-SMA. Moreover, CEPO significantly promoted tubular epithelial cell proliferation without erythropoiesis. In conclusion, we identified a new therapeutic approach using CEPO to protect kidneys from ischemia-reperfusion injury.

  10. Sustained release formulation of erythropoietin using hyaluronic acid hydrogels crosslinked by Michael addition.

    PubMed

    Hahn, Sei Kwang; Oh, Eun Ju; Miyamoto, Hajime; Shimobouji, Tsuyoshi

    2006-09-28

    A novel sustained release formulation of erythropoietin (EPO) was successfully developed using hyaluronic acid (HA) hydrogels crosslinked by Michael addition. Adipic acid dihydrazide grafted HA (HA-ADH) was prepared and then modified into methacrylated HA (HA-MA). (1)H NMR analysis showed that the degrees of HA-ADH and HA-MA modification were 69 and 29 mol%, respectively. Using the specific crosslinkers of dithiothreitol (DTT) and peptide linker, EPO was loaded during HA-MA hydrogel preparation by Michael addition chemistry between thiol and methacrylate groups. The amount of EPO recovered from both hydrogels after degradation with hyaluronidase SD (HAse SD) was about 90%. The crosslinking reaction with peptide linker (GCYKNRDCG) was faster than that with DTT. The gelation time was about 30 min for peptide linker and 180 min for DTT. In vitro release test of EPO from HA-MA hydrogel at 37 degrees C showed that EPO was released rapidly for 2 days and then slowly up to 7 days from HA-MA hydrogels. The released EPO appeared to be intact from the analysis with RP-HPLC. According to in vivo release test of EPO from HA-MA hydrogels crosslinked with the peptide linker in Sprague-Dawley (SD) rats, elevated plasma concentration of EPO was maintained up to 7 days. There was no adverse effect during and after the in vivo tests. PMID:16781096

  11. Mass spectrometry-based glycoproteomic approach involving lysine derivatization for structural characterization of recombinant human erythropoietin.

    PubMed

    Cindrić, Mario; Bindila, Laura; Cepo, Tina; Peter-Katalinić, Jasna

    2006-11-01

    Lysine-containing peptides comprising glycosylation sites derived from recombinant human erythropoietin (rHuEPO) by trypsin or Lys-C and PNGase F dual digestion were derivatized with 2-methoxy-4,5-dihydro-1H-imidazole and its deuterated analogues. In the same reaction, under reducing conditions (beta-mercaptoethanol), cysteines were converted into methyl-cysteines and lysines into Lys-4,5-dihydro-1H-imidazole. Both modifications on cysteines and lysines simplified the CID-MS/MS spectra, while preserving the structural information by yielding y-series ions and improved the mass spectral signal intensity up to 25 times. Moreover, by this approach, the N-glycan occupation sites were unambiguously determined. O-Glycosylation sites as well as O-glycan structures were determined by a LC-MS/MS experiment carried out on dually digested rHuEPO. N-Glycan mixture purified on a graphitized carbon column using a newly developed method that extracted only sialylated carbohydrates was analyzed first using MALDI-TOF in negative linear ion mode with low mass accuracy but without interferences and metastabile ions and then a reflectron with high mass accuracy. After defining the precursor ions, we performed the nanoESI QTOF MS/MS analysis on N-glycans, mainly targeting the distinction between carbohydrates with sialylated antennae and those lacking sialic acid moieties.

  12. Renal epithelium regulates erythropoiesis via HIF-dependent suppression of erythropoietin

    PubMed Central

    Farsijani, Navid M.; Liu, Qingdu; Davidoff, Olena; Sha, Feng; Fandrey, Joachim; Ikizler, T. Alp; O’Connor, Paul M.

    2016-01-01

    The adult kidney plays a central role in erythropoiesis and is the main source of erythropoietin (EPO), an oxygen-sensitive glycoprotein that is essential for red blood cell production. Decreases of renal pO2 promote hypoxia-inducible factor 2–mediated (HIF-2–mediated) induction of EPO in peritubular interstitial fibroblast-like cells, which serve as the cellular site of EPO synthesis in the kidney. It is not clear whether HIF signaling in other renal cell types also contributes to the regulation of EPO production. Here, we used a genetic approach in mice to investigate the role of renal epithelial HIF in erythropoiesis. Specifically, we found that HIF activation in the proximal nephron via induced inactivation of the von Hippel–Lindau tumor suppressor, which targets the HIF-α subunit for proteasomal degradation, led to rapid development of hypoproliferative anemia that was associated with a reduction in the number of EPO-producing renal interstitial cells. Moreover, suppression of renal EPO production was associated with increased glucose uptake, enhanced glycolysis, reduced mitochondrial mass, diminished O2 consumption, and elevated renal tissue pO2. Our genetic analysis suggests that tubulointerstitial cellular crosstalk modulates renal EPO production under conditions of epithelial HIF activation in the kidney. PMID:26927670

  13. Human Erythropoietin Gene Delivery Using an Arginine-grafted Bioreducible Polymer System

    PubMed Central

    Lee, Youngsook; Nam, Hye Yeong; Kim, Jaesung; Lee, Minhyung; Yockman, James W; Shin, Sug Kyun; Kim, Sung Wan

    2012-01-01

    Erythropoiesis-stimulating agents are widely used to treat anemia for chronic kidney disease (CKD) and cancer, however, several clinical limitations impede their effectiveness. Nonviral gene therapy systems are a novel solution to these problems as they provide stable and low immunogenic protein expression levels. Here, we show the application of an arginine-grafted bioreducible poly(disulfide amine) (ABP) polymer gene delivery system as a platform for in vivo transfer of human erythropoietin plasmid DNA (phEPO) to produce long-term, therapeutic erythropoiesis. A single systemic injection of phEPO/ABP polyplex led to higher hematocrit levels over a 60-day period accompanied with reticulocytosis and high hEPO protein expression. In addition, we found that the distinct temporal and spatial distribution of phEPO/ABP polyplexes contributed to increased erythropoietic effects compared to those of traditional EPO therapies. Overall, our study suggests that ABP polymer-based gene therapy provides a promising clinical strategy to reach effective therapeutic levels of hEPO gene. PMID:22472948

  14. Measurement of sialic acid content is insufficient to assess bioactivity of recombinant human erythropoietin.

    PubMed

    Yanagihara, Shigehiro; Taniguchi, Yuya; Hosono, Mareto; Yoshioka, Eiji; Ishikawa, Rika; Shimada, Yoshihiro; Kadoya, Toshihiko; Kutsukake, Kazuhiro

    2010-01-01

    Assessment of biological potency and its comparison with clinical effects are important in the quality control of therapeutic glycoproteins. Animal models are usually used for evaluating bioactivity of these compounds. However, alternative methods are required to simplify the bioassay and avoid ethical issues associated with animal studies. Negatively charged sialic acid residues are known to be critical for in vivo bioactivity of recombinant human erythropoietin (rhEPO). In this study, we used capillary zone electrophoresis, a charge-based separation method, to estimate the sialic acid content for predicting in vivo bioactivity of rhEPO. In vivo bioactivities of rhEPO subfractions were measured and compared with sialylation levels. The results obtained indicated that in vivo bioactivity of rhEPO is not simply correlated with the sialylation level, which suggests that it is difficult to predict biological potency from the sialic acid content alone. N-Glycan moieties as well as sialic acid residues may have a significant impact on in vivo bioactivity of rhEPO. PMID:20823580

  15. Resistance to Recombinant Human Erythropoietin Therapy in a Rat Model of Chronic Kidney Disease Associated Anemia

    PubMed Central

    Garrido, Patrícia; Ribeiro, Sandra; Fernandes, João; Vala, Helena; Rocha-Pereira, Petronila; Bronze-da-Rocha, Elsa; Belo, Luís; Costa, Elísio; Santos-Silva, Alice; Reis, Flávio

    2015-01-01

    This study aimed to elucidate the mechanisms explaining the persistence of anemia and resistance to recombinant human erythropoietin (rHuEPO) therapy in a rat model of chronic kidney disease (CKD)-associated anemia with formation of anti-rHuEPO antibodies. The remnant kidney rat model of CKD induced by 5/6 nephrectomy was used to test a long-term (nine weeks) high dose of rHuEPO (200 UI/kg bw/week) treatment. Hematological and biochemical parameters were evaluated as well as serum and tissue (kidney, liver and/or duodenum) protein and/or gene expression of mediators of erythropoiesis, iron metabolism and tissue hypoxia, inflammation, and fibrosis. Long-term treatment with a high rHuEPO dose is associated with development of resistance to therapy as a result of antibodies formation. In this condition, serum EPO levels are not deficient and iron availability is recovered by increased duodenal absorption. However, erythropoiesis is not stimulated, and the resistance to endogenous EPO effect and to rHuEPO therapy results from the development of a hypoxic, inflammatory and fibrotic milieu in the kidney tissue. This study provides new insights that could be important to ameliorate the current therapeutic strategies used to treat patients with CKD-associated anemia, in particular those that become resistant to rHuEPO therapy. PMID:26712750

  16. Cell surface activation of the erythropoietin receptor by Friend spleen focus-forming virus gp55.

    PubMed Central

    Li, J P; Hu, H O; Niu, Q T; Fang, C

    1995-01-01

    The leukemogenic membrane glycoprotein gp55, encoded by Friend spleen focus-forming virus (SFFV), induces erythroid cell proliferation through its interaction with the erythropoietin receptor (EPO-R). There are two forms of gp55 in SFFV-infected cells: an intracellular form (more than 95% of the total protein), which is localized within the endoplasmic reticulum (ER) membranes, and a cell surface form (about 3 to 5%). Because both forms of the viral proteins bind to EPO-R, it is not clear whether the viral protein induces mitogenesis intracellularly or at the cell surface. To address this question, we constructed an EPO-R mutant that contained a 6-amino-acid (DEKKMP) C-terminus ER retention signal. Biochemical and functional analyses with this mutant indicated that it was completely retained in the ER and not expressed at the cell surface. Further analysis showed that the mutant, like the wild-type EPO-R, interacted with SFFV gp55. However, this apparent intracellular interaction between the two proteins failed to induce growth factor-independent proliferation of Ba/F3 cells. Furthermore, spontaneous variants of the ER-retained EPO-R selected on the basis of their ability to induce cell proliferation when coexpressed with gp55 were exclusively expressed at the cell surface. Thus, our results support the hypothesis that the mitogenic activation of the EPO-R by gp55 requires the interaction of the two proteins at the cell surface. PMID:7853508

  17. VEGF-mediated STAT3 activation inhibits retinal vascularization by down-regulating local erythropoietin expression.

    PubMed

    Wang, Haibo; Byfield, Grace; Jiang, Yanchao; Smith, George Wesley; McCloskey, Manabu; Hartnett, M Elizabeth

    2012-03-01

    Avascular, hypoxic retina has been postulated to be a source of angiogenic factors that cause aberrant angiogenesis and intravitreal neovascularization (IVNV) in retinopathy of prematurity. Vascular endothelial growth factor (VEGF) is an important factor involved. However, VEGF is also required for normal retinal vascular development, which raises concerns about inhibiting its activity to treat IVNV in retinopathy of prematurity. Therefore, understanding the effects that VEGF has on other factors in the development of avascular retina is important to prevent aberrant angiogenesis and IVNV. Here, we show that STAT3 was activated by increased retinal VEGF in the rat 50/10 oxygen-induced retinopathy model. Phospho-STAT3 colocalized with glutamine synthetase-labeled Müller cells. Inhibition of STAT3 reduced avascular retina and increased retinal erythropoietin (Epo) expression. Epo administered exogenously also reduced avascular retina in the model. In an in vitro study, hypoxia-induced VEGF inhibited Epo gene expression by STAT3 activation in rat Müller cells. The mechanism by which activated STAT3 regulated Epo was by inhibition of Epo promoter activity. Together, these findings show that increased retinal VEGF contributes to avascular retina by regulating retinal Epo expression through Janus kinase/STAT signaling. Our results suggest that rescuing Epo expression in the retina before the development of IVNV may promote normal developmental angiogenesis and, therefore, reduce the stimulus for later pathologic IVNV.

  18. Inhibitory effects of tetradecanoylphorbol acetate and diacylglycerol on erythropoietin production in human renal carcinoma cell cultures

    SciTech Connect

    Hagiwara, Masamichi; Nagakura, Kazuhiko; Ueno, Munehisa; Fisher, J.W. )

    1987-11-01

    A human renal carcinoma from a patient with an erythrocytosis, serially transplanted into athymic nude mice, was grown in primary monolayer cell cultures. After reaching confluency the cultured cells formed multicellular hemicysts (domes) which became more abundant as the cultures approached saturation density. Erythropoietin (Ep) production by this renal carcinoma in culture was only slightly increased at the time of semiconfluency but showed a marked increase in Ep levels in the culture medium after the cultures reached confluency, in parallel with an increase in dome formation. The phorbol ester tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) showed a significant dose-related inhibitory effect on Ep production and dome formation in the renal carcinoma cell cultures, suggesting an important role of protein kinase C, the only known receptor for TPA, in inhibiting the expression of differentiated phenotypes in the renal carcinoma cells. These studies suggest a role of the inositol-lipid second messenger path and protein kinase C in the regulation of Ep production.

  19. Molecular and functional characterization of erythropoietin receptor of the goldfish (Carassius auratus L.).

    PubMed

    Katakura, Fumihiko; Katzenback, Barbara A; Belosevic, Miodrag

    2014-08-01

    Erythropoietin receptor (EPOR) is a member of the class I cytokine receptor superfamily and signaling through this receptor is important for the proliferation, differentiation and survival of erythrocyte progenitor cells. This study reports on the molecular and functional characterization of goldfish EPOR. The identified goldfish EPOR sequence possesses the conserved EPOR ligand binding domain, the fibronectin domain, the class I cytokine receptor superfamily motif (WSXWS) as well as several intracellular signaling motifs characteristic of other vertebrate EPORs. The expression of epor mRNA in goldfish tissues, cell populations and cells treated with recombinant goldfish EPO (rgEPO) were evaluated by quantitative PCR revealing that goldfish epor mRNA is transcribed in both erythropoietic tissues (blood, kidney and spleen) and non-hematopoietic tissues (brain, heart and gill), as well as in immature erythrocytes. Recombinant goldfish EPOR (rgEPOR), consisting of its extracellular domain, dose-dependently inhibited proliferation of progenitor cells induced by rgEPO. In vitro binding studies indicated that rgEPO exists as monomer, dimer and/or trimmer and that rgEPOR exists as monomer and/or homodimer, and when incubated together, formed a ligand-receptor complex. Our results demonstrate that goldfish EPO/EPOR signaling has been highly conserved throughout vertebrate evolution as a required mechanism for erythrocyte development.

  20. Renal epithelium regulates erythropoiesis via HIF-dependent suppression of erythropoietin.

    PubMed

    Farsijani, Navid M; Liu, Qingdu; Kobayashi, Hanako; Davidoff, Olena; Sha, Feng; Fandrey, Joachim; Ikizler, T Alp; O'Connor, Paul M; Haase, Volker H

    2016-04-01

    The adult kidney plays a central role in erythropoiesis and is the main source of erythropoietin (EPO), an oxygen-sensitive glycoprotein that is essential for red blood cell production. Decreases of renal pO2 promote hypoxia-inducible factor 2-mediated (HIF-2-mediated) induction of EPO in peritubular interstitial fibroblast-like cells, which serve as the cellular site of EPO synthesis in the kidney. It is not clear whether HIF signaling in other renal cell types also contributes to the regulation of EPO production. Here, we used a genetic approach in mice to investigate the role of renal epithelial HIF in erythropoiesis. Specifically, we found that HIF activation in the proximal nephron via induced inactivation of the von Hippel-Lindau tumor suppressor, which targets the HIF-α subunit for proteasomal degradation, led to rapid development of hypoproliferative anemia that was associated with a reduction in the number of EPO-producing renal interstitial cells. Moreover, suppression of renal EPO production was associated with increased glucose uptake, enhanced glycolysis, reduced mitochondrial mass, diminished O2 consumption, and elevated renal tissue pO2. Our genetic analysis suggests that tubulointerstitial cellular crosstalk modulates renal EPO production under conditions of epithelial HIF activation in the kidney. PMID:26927670

  1. Erythropoietin production in neuroepithelial and neural crest cells during primitive erythropoiesis.

    PubMed

    Suzuki, Norio; Hirano, Ikuo; Pan, Xiaoqing; Minegishi, Naoko; Yamamoto, Masayuki

    2013-01-01

    Erythropoietin (Epo) supports both primitive erythropoiesis in the yolk sac and definitive erythropoiesis in the fetal liver and bone marrow. Although definitive erythropoiesis requires kidney- and liver-secreted Epo, it is unclear which cells produce Epo for primitive erythropoiesis. Here we find neural Epo-producing (NEP) cells in mid-gestational stage embryos using mouse lines that express green fluorescent protein (GFP) under the Epo gene regulation. In these mice, GFP is expressed exclusively in a subpopulation of neural and neural crest cells at embryonic day 9.0 when Epo-deficient embryos exhibit abnormalities in primitive erythropoiesis. The GFP-positive NEP cells express Epo mRNA and the ex vivo culture of embryonic day 8.5 neural tubes results in the secretion of Epo, which is able to induce the proliferation and differentiation of yolk sac-derived erythroid cells. These results thus suggest that NEP cells secrete Epo and might support the development of primitive erythropoiesis. PMID:24309470

  2. Roles of renal erythropoietin-producing (REP) cells in the maintenance of systemic oxygen homeostasis.

    PubMed

    Suzuki, Norio; Yamamoto, Masayuki

    2016-01-01

    Erythropoietic induction is critical for enhancing the efficiency of oxygen delivery during the chronic phase of the systemic hypoxia response. The erythroid growth factor erythropoietin (Epo) triggers the erythropoietic induction through the activation of erythroid genes related to cell survival, differentiation, and iron metabolism. Because Epo is produced in renal Epo-producing (REP) cells in a hypoxia-inducible manner, REP cells serve as a control center for the systemic hypoxia response. In fact, the loss of Epo production in REP cells causes chronic severe anemia in genetically modified mice, and REP cell-specific inactivation of PHD2 (prolyl-hydroxylase domain enzyme 2) results in erythrocytosis via overexpression of the Epo gene due to the constitutive activation of HIF2α (hypoxia-inducible transcription factor 2α). REP cells are located in the interstitial spaces between renal tubules and capillaries, where the oxygen supply is low but oxygen consumption is high, for the highly sensitive detection of decreased oxygen supplies to the body. Under disease conditions, REP cells transform to myofibroblasts and lose their Epo-producing ability. Therefore, elucidation of Epo gene regulation and REP cell features directly contributes to understanding the pathology of chronic kidney disease. To further analyze REP cells, we introduce a newly established mouse line in which REP cells are efficiently labeled with fluorescent protein. PMID:26452589

  3. Progress in detecting cell-surface protein receptors: the erythropoietin receptor example.

    PubMed

    Elliott, Steve; Sinclair, Angus; Collins, Helen; Rice, Linda; Jelkmann, Wolfgang

    2014-02-01

    Testing for the presence of specific cell-surface receptors (such as EGFR or HER2) on tumor cells is an integral part of cancer care in terms of treatment decisions and prognosis. Understanding the strengths and limitations of these tests is important because inaccurate results may occur if procedures designed to prevent false-negative or false-positive outcomes are not employed. This review discusses tests commonly used to identify and characterize cell-surface receptors, such as the erythropoietin receptor (EpoR). First, a summary is provided on the biology of the Epo/EpoR system, describing how EpoR is expressed on erythrocytic progenitors and precursors in the bone marrow where it mediates red blood cell production in response to Epo. Second, studies are described that investigated whether erythropoiesis-stimulating agents could stimulate tumor progression in cancer patients and whether EpoR is expressed and functional on tumor cells or on endothelial cells. The methods used in these studies included immunohistochemistry, Northern blotting, Western blotting, and binding assays. This review summarizes the strengths and limitations of these methods. Critically analyzing data from tests for cell-surface receptors such as EpoR requires understanding the techniques utilized and demonstrating that results are consistent with current knowledge about receptor biology. PMID:24337485

  4. Erythropoietin gene expression: developmental-stage specificity, cell-type specificity, and hypoxia inducibility.

    PubMed

    Suzuki, Norio

    2015-01-01

    Erythrocytes play an essential role in the delivery of oxygen from the lung to every organ; a decrease in erythrocytes (anemia) causes hypoxic stress and tissue damage. To maintain oxygen homeostasis in adult mammals, when the kidney senses hypoxia, it secretes an erythroid growth factor, erythropoietin (Epo), which stimulates erythropoiesis in the bone marrow. Recently, studies using genetically modified mice have shown that the in vivo expression profile of the Epo gene changes dramatically during development. The first Epo-producing cells emerge in the neural crest and neuroepithelium of mid-stage embryos and support primitive erythropoiesis in the yolk sac. Subsequently, Epo from the hepatocytes stimulates erythropoiesis in the fetal liver of later stage embryos in a paracrine manner. In fact, erythroid lineage cells comprise the largest cell population in the fetal liver, and hepatocytes are distributed among the erythroid cell clusters. Adult erythropoiesis in the bone marrow requires Epo that is secreted by renal Epo-producing cells (REP cells). REP cells are widely distributed in the renal cortex and outer medulla. Hypoxia-inducible Epo production both in hepatocytes and REP cells is controlled at the gene transcription level that is mainly mediated by the hypoxia-inducible transcription factor (HIF) pathway. These mouse studies further provide insights into the molecular mechanisms of the cell-type specific, hypoxia-inducible expression of the Epo gene, which involves multiple sets of cis- and trans-regulatory elements. PMID:25786542

  5. Midazolam inhibits the hypoxia-induced up-regulation of erythropoietin in the central nervous system.

    PubMed

    Matsuyama, Tomonori; Tanaka, Tomoharu; Tatsumi, Kenichiro; Daijo, Hiroki; Kai, Shinichi; Harada, Hiroshi; Fukuda, Kazuhiko

    2015-08-15

    Erythropoietin (EPO), a regulator of red blood cell production, is endogenously expressed in the central nervous system. It is mainly produced by astrocytes under hypoxic conditions and has proven to have neuroprotective and neurotrophic effects. In the present study, we investigated the effect of midazolam on EPO expression in primary cultured astrocytes and the mouse brain. Midazolam was administered to 6-week-old BALB/c male mice under hypoxic conditions and pregnant C57BL/6N mice under normoxic conditions. Primary cultured astrocytes were also treated with midazolam under hypoxic conditions. The expression of EPO mRNA in mice brains and cultured astrocytes was studied. In addition, the expression of hypoxia-inducible factor (HIF), known as the main regulator of EPO, was evaluated. Midazolam significantly reduced the hypoxia-induced up-regulation of EPO in BALB/c mice brains and primary cultured astrocytes and suppressed EPO expression in the fetal brain. Midazolam did not affect the total amount of HIF proteins but significantly inhibited the nuclear expression of HIF-1α and HIF-2α proteins. These results demonstrated the suppressive effects of midazolam on the hypoxia-induced up-regulation of EPO both in vivo and in vitro. PMID:26001375

  6. Erythropoietin Exacerbates Inflammation and Increases the Mortality of Histoplasma capsulatum-Infected Mice.

    PubMed

    Locachevic, Gisele Aparecida; Pereira, Priscilla Aparecida Tartari; Secatto, Adriana; Fontanari, Caroline; Galvão, Alyne Fávero; Prado, Morgana Kelly Borges; Zoccal, Karina Furlani; Petta, Tânia; Moraes, Luiz Alberto Beraldo; Ramos, Simone Gusmão; de Castro, Fabíola Attie; Sorgi, Carlos Artério; Faccioli, Lúcia Helena

    2015-01-01

    Erythropoietin (EPO) is a key hormone involved in red blood cell formation, but its effects on nonerythroid cells, such as macrophages, have not been described. Macrophages are key cells in controlling histoplasmosis, a fungal infection caused by Histoplasma capsulatum (Hc). Considering that little is known about EPO's role during fungal infections and its capacity to activate macrophages, in this study we investigated the impact of EPO pretreatment on the alveolar immune response during Hc infection. The consequence of EPO pretreatment on fungal infection was determined by evaluating animal survival, fungal burden, activation of bronchoalveolar macrophages, inflammatory mediator release, and lung inflammation. Pretreatment with EPO diminished mononuclear cell numbers, increased the recruitment of F4/80(+)/CD80(+) and F4/80(+)/CD86(+) cells to the bronchoalveolar space, induced higher production of IFN-γ, IL-6, MIP-1α, MCP-1, and LTB4, reduced PGE2 concentration, and did not affect fungal burden. As a consequence, we observed an increase in lung inflammation with extensive tissue damage that might account for augmented mouse mortality after infection. Our results demonstrate for the first time that EPO treatment has a deleterious impact on lung immune responses during fungal infection. PMID:26538835

  7. Erythropoietin is a hypoxia inducible factor-induced protective molecule in experimental autoimmune neuritis.

    PubMed

    Luo, Bangwei; Jiang, Man; Yang, Xiaofeng; Zhang, Zhiyuan; Xiong, Jian; Schluesener, Hermann J; Zhang, Zhiren; Wu, Yuzhang

    2013-08-01

    Experimental autoimmune neuritis (EAN), an autoantigen-specific T-cell-mediated disease model for human demyelinating inflammatory disease of the peripheral nervous system, is characterized by self-limitation. Here we investigated the regulation and contribution of erythropoietin (EPO) in EAN self-limitation. In EAN sciatic nerves, hypoxia, and protein and mRNA levels of hypoxia-inducible factor 1α (HIF-1α), HIF-2α, EPO and EPO receptor (EPOR) were induced in parallel at disease peak phase but reduced at recovery periods. Further, the deactivation of HIF reduced EAN-induced EPO/EPOR upregulation in EAN, suggesting the central contribution of HIF to EPO/EPOR induction. The deactivation of EPOR signalling exacerbated EAN progression, implying that endogenous EPO contributed to EAN recovery. Exogenous EPO treatment greatly improved EAN recovery. In addition, EPO was shown to promote Schwann cell survival and myelin production. In EAN, EPO treatment inhibited lymphocyte proliferation and altered helper T cell differentiation by inducing increase of Foxp3(+)/CD4(+) regulatory T cells and decrease of IFN-γ(+)/CD4(+) Th1 cells. Furthermore, EPO inhibited inflammatory macrophage activation and promoted its phagocytic activity. In summary, our data demonstrated that EPO was induced in EAN by HIF and contributed to EAN recovery, and endogenous and exogenous EPO could effectively suppress EAN by attenuating inflammation and exerting direct cell protection, indicating that EPO contributes to the self-recovery of EAN and could be a potent candidate for treatment of autoimmune neuropathies. PMID:23603807

  8. Distinct subpopulations of FOXD1 stroma-derived cells regulate renal erythropoietin.

    PubMed

    Kobayashi, Hanako; Liu, Qingdu; Binns, Thomas C; Urrutia, Andres A; Davidoff, Olena; Kapitsinou, Pinelopi P; Pfaff, Andrew S; Olauson, Hannes; Wernerson, Annika; Fogo, Agnes B; Fong, Guo-Hua; Gross, Kenneth W; Haase, Volker H

    2016-05-01

    Renal peritubular interstitial fibroblast-like cells are critical for adult erythropoiesis, as they are the main source of erythropoietin (EPO). Hypoxia-inducible factor 2 (HIF-2) controls EPO synthesis in the kidney and liver and is regulated by prolyl-4-hydroxylase domain (PHD) dioxygenases PHD1, PHD2, and PHD3, which function as cellular oxygen sensors. Renal interstitial cells with EPO-producing capacity are poorly characterized, and the role of the PHD/HIF-2 axis in renal EPO-producing cell (REPC) plasticity is unclear. Here we targeted the PHD/HIF-2/EPO axis in FOXD1 stroma-derived renal interstitial cells and examined the role of individual PHDs in REPC pool size regulation and renal EPO output. Renal interstitial cells with EPO-producing capacity were entirely derived from FOXD1-expressing stroma, and Phd2 inactivation alone induced renal Epo in a limited number of renal interstitial cells. EPO induction was submaximal, as hypoxia or pharmacologic PHD inhibition further increased the REPC fraction among Phd2-/- renal interstitial cells. Moreover, Phd1 and Phd3 were differentially expressed in renal interstitium, and heterozygous deficiency for Phd1 and Phd3 increased REPC numbers in Phd2-/- mice. We propose that FOXD1 lineage renal interstitial cells consist of distinct subpopulations that differ in their responsiveness to Phd2 inactivation and thus regulation of HIF-2 activity and EPO production under hypoxia or conditions of pharmacologic or genetic PHD inactivation. PMID:27088801

  9. Erythropoietin Exacerbates Inflammation and Increases the Mortality of Histoplasma capsulatum-Infected Mice

    PubMed Central

    Locachevic, Gisele Aparecida; Pereira, Priscilla Aparecida Tartari; Secatto, Adriana; Fontanari, Caroline; Galvão, Alyne Fávero; Prado, Morgana Kelly Borges; Zoccal, Karina Furlani; Petta, Tânia; Moraes, Luiz Alberto Beraldo; Ramos, Simone Gusmão; de Castro, Fabíola Attie; Sorgi, Carlos Artério; Faccioli, Lúcia Helena

    2015-01-01

    Erythropoietin (EPO) is a key hormone involved in red blood cell formation, but its effects on nonerythroid cells, such as macrophages, have not been described. Macrophages are key cells in controlling histoplasmosis, a fungal infection caused by Histoplasma capsulatum (Hc). Considering that little is known about EPO's role during fungal infections and its capacity to activate macrophages, in this study we investigated the impact of EPO pretreatment on the alveolar immune response during Hc infection. The consequence of EPO pretreatment on fungal infection was determined by evaluating animal survival, fungal burden, activation of bronchoalveolar macrophages, inflammatory mediator release, and lung inflammation. Pretreatment with EPO diminished mononuclear cell numbers, increased the recruitment of F4/80+/CD80+ and F4/80+/CD86+ cells to the bronchoalveolar space, induced higher production of IFN-γ, IL-6, MIP-1α, MCP-1, and LTB4, reduced PGE2 concentration, and did not affect fungal burden. As a consequence, we observed an increase in lung inflammation with extensive tissue damage that might account for augmented mouse mortality after infection. Our results demonstrate for the first time that EPO treatment has a deleterious impact on lung immune responses during fungal infection. PMID:26538835

  10. Erythropoietin (EPO) increases myelin gene expression in CG4 oligodendrocyte cells through the classical EPO receptor.

    PubMed

    Cervellini, Ilaria; Annenkov, Alexander; Brenton, Thomas; Chernajovsky, Yuti; Ghezzi, Pietro; Mengozzi, Manuela

    2013-08-28

    Erythropoietin (EPO) has protective effects in neurodegenerative and neuroinflammatory diseases, including in animal models of multiple sclerosis, where EPO decreases disease severity. EPO also promotes neurogenesis and is protective in models of toxic demyelination. In this study, we asked whether EPO could promote neurorepair by also inducing remyelination. In addition, we investigated whether the effect of EPO could be mediated by the classical erythropoietic EPO receptor (EPOR), since it is still questioned if EPOR is functional in nonhematopoietic cells. Using CG4 cells, a line of rat oligodendrocyte precursor cells, we found that EPO increases the expression of myelin genes (myelin oligodendrocyte glycoprotein [MOG] and myelin basic protein [MBP]). EPO had no effect in wild-type CG4 cells, which do not express EPOR, whereas it increased MOG and MBP expression in cells engineered to overexpress EPOR (CG4-EPOR). This was reflected in a marked increase in MOG protein levels, as detected by Western blot. In these cells, EPO induced by 10-fold the early growth response gene 2 (Egr2), which is required for peripheral myelination. However, Egr2 silencing with a siRNA did not reverse the effect of EPO, indicating that EPO acts through other pathways. In conclusion, EPO induces the expression of myelin genes in oligodendrocytes and this effect requires the presence of EPOR. This study demonstrates that EPOR can mediate neuroreparative effects.

  11. Salmonella Infection Enhances Erythropoietin Production by the Kidney and Liver, Which Correlates with Elevated Bacterial Burdens.

    PubMed

    Li, Lin-Xi; Benoun, Joseph M; Weiskopf, Kipp; Garcia, K Christopher; McSorley, Stephen J

    2016-10-01

    Salmonella infection profoundly affects host erythroid development, but the mechanisms responsible for this effect remain poorly understood. We monitored the impact of Salmonella infection on erythroid development and found that systemic infection induced anemia, splenomegaly, elevated erythropoietin (EPO) levels, and extramedullary erythropoiesis in a process independent of Salmonella pathogenicity island 2 (SPI2) or flagellin. The circulating EPO level was also constitutively higher in mice lacking the expression of signal-regulatory protein α (SIRPα). The expression level of EPO mRNA was elevated in the kidney and liver but not increased in the spleens of infected mice despite the presence of extramedullary erythropoiesis in this tissue. In contrast to data from a previous report, mice lacking EPO receptor (EPOR) expression on nonerythroid cells (EPOR rescued) had bacterial loads similar to those of wild-type mice following Salmonella infection. Indeed, treatment to reduce splenic erythroblasts and mature red blood cells correlated with elevated bacterial burdens, implying that extramedullary erythropoiesis benefits the host. Together, these findings emphasize the profound effect of Salmonella infection on erythroid development and suggest that the modulation of erythroid development has both positive and negative consequences for host immunity.

  12. Erythropoietin Slows Photoreceptor Cell Death in a Mouse Model of Autosomal Dominant Retinitis Pigmentosa

    PubMed Central

    Kasmala, Lorraine; Bond, Wesley S.; de Lucas Cerrillo, Ana M.; Wynn, Kristi; Lewin, Alfred S.

    2016-01-01

    Purpose To test the efficacy of systemic gene delivery of a mutant form of erythropoietin (EPO-R76E) that has attenuated erythropoietic activity, in a mouse model of autosomal dominant retinitis pigmentosa. Methods Ten-day old mice carrying one copy of human rhodopsin with the P23H mutation and both copies of wild-type mouse rhodopsin (hP23H RHO+/-,mRHO+/+) were injected into the quadriceps with recombinant adeno-associated virus (rAAV) carrying either enhanced green fluorescent protein (eGFP) or EpoR76E. Visual function (electroretinogram) and retina structure (optical coherence tomography, histology, and immunohistochemistry) were assessed at 7 and 12 months of age. Results The outer nuclear layer thickness decreased over time at a slower rate in rAAV.EpoR76E treated as compared to the rAAV.eGFP injected mice. There was a statistically significant preservation of the electroretinogram at 7, but not 12 months of age. Conclusions Systemic EPO-R76E slows death of the photoreceptors and vision loss in hP23H RHO+/-,mRHO+/+ mice. Treatment with EPO-R76E may widen the therapeutic window for retinal degeneration patients by increasing the number of viable cells. Future studies might investigate if co-treatment with EPO-R76E and gene replacement therapy is more effective than gene replacement therapy alone. PMID:27299810

  13. Interleukin 3 promotes erythroid burst formation in serum-free cultures without detectable erythropoietin

    SciTech Connect

    Goodman, J.W.; Hall, E.A.; Miller, K.L.; Shinpock, S.G.

    1985-05-01

    Erythroid burst-forming units (BFU-E) from mouse bone marrow were grown for 7 days in agar or serum-free methylcellulose cultures in the presence or absence of erythropoietin (Ep) and/or interleukin 3 (IL-3). It was found that IL-3, even in the absence of serum and detectable Ep, was able to stimulate the full development of many erythroid bursts. This IL-3 effect was cell-dose dependent and did not appear to correlate with Ep dose. Spontaneous bursts and those stimulated by Ep only were rare and when seen were very small relative to those produced by IL-3 or IL-3 plus Ep. When addition of IL-3 or Ep to 7-day cultures was delayed, IL-3 but not Ep was shown to maintain BFU-E. No evidence was found by radioimmunoassay that Ep was produced or released in 7-day, serum-free cultures of bone marrow nor was Ep activity detected in culture media except those to which it had been added deliberately.

  14. A/sub 1/ and A/sub 2/ adenosine receptor regulation of erythropoietin production

    SciTech Connect

    Ueno, M.; Brookins, J.; Beckman, B.; Fisher, J.W.

    1988-01-01

    The effects of adenosine (ADE) and ADE agonists on erythropoietin (Ep) production were determined using percent (%) /sup 59/Fe incorporation in red cells of exhypoxic polycythemic mice. The hemisulfate salt of ADE produced a significant increase in % /sup 59/Fe incorporation in response to hypoxia in concentrations of 400 to 1600 nmol/kg/day. 5'-N-ethyl-carboxamideadenosine (NECA), a selective A/sub 2/ receptor agonist, increased radioiron incorporation in a dose-dependent manner. In contrast, N/sup 6/-cyclohexyladenosine (CHA), a selective A/sub 1/ receptor agonist, did not affect radioiron incorporation in concentrations up to 1600 nmol/kg/day. Albuterol, a beta 2-adrenergic agonist, enhanced % /sup 59/Fe incorporation in polycythemic mice and low doses of CHA, which were not effective alone on % /sup 59/Fe incorporation in polycythemic mice exposed to hypoxia, inhibited the enhancement in radioiron induced by albuterol plus hypoxia. Theophylline, a well-known antagonist of ADE receptors, blocked the ADE and NECA enhancement in radioiron incorporation at a dose of theophylline alone which produced only a slight enhancement of % /sup 59/Fe incorporation.

  15. Antidepressant-Like Effects of Erythropoietin: A Focus on Behavioural and Hippocampal Processes

    PubMed Central

    Osborn, Meagan; Rustom, Nazneen; Clarke, Melanie; Litteljohn, Darcy; Rudyk, Chris; Anisman, Hymie; Hayley, Shawn

    2013-01-01

    Depression is a chronic and debilitating condition with a significant degree of relapse and treatment resistance that could stem, at least in part, from disturbances of neuroplasticity. This has led to an increased focus on treatment strategies that target brain derived neurotrophic factor (BDNF), synaptic plasticity and adult neurogenesis. In the current study we aimed to assess whether erythropoietin (EPO) would have antidepressant-like effects given its already established pro-trophic actions. In particular, we assessed whether EPO would diminish the deleterious effects of a social stressor in mice. Indeed, EPO induced anxiolytic and antidepressant-like responses in a forced swim test, open field, elevated-plus maze, and a novelty test, and appeared to blunt some of the negative behavioural effects of a social stressor. Furthermore, EPO promoted adult hippocampal neurogenesis, an important feature of effective antidepressants. Finally, a separate study using the mTOR inhibitor rapamycin revealed that antagonizing this pathway prevented the impact of EPO upon forced swim performance. These data are consistent with previous findings showing that the mTOR pathway and its neurogenic and synaptogenic effects might mediate the behavioral consequences of antidepressant agents. Our findings further highlight EPO as a possible adjunct treatment for affective disorders, as well as other stressor associated disorders of impaired neuroplasticity. PMID:24019878

  16. Recombinant human erythropoietin (rHuEPO): more than just the correction of uremic anemia.

    PubMed

    Buemi, Michele; Aloisi, Carmela; Cavallaro, Emanuela; Corica, Francesco; Floccari, Fulvio; Grasso, Giovanni; Lasco, Antonino; Pettinato, Giuseppina; Ruello, Antonella; Sturiale, Alessio; Frisina, Nicola

    2002-01-01

    Hematopoiesis is controlled by numerous interdependent humoral and endocrine factors. Erythropoietin (EPO), a hydrophobic sialoglycoproteic hormone, plays a crucial role in the regulation of hematopoiesis, and induces proliferation, maturation and differentiation of the erythroid cell line precursors. Thanks to recombinant DNA techniques, different recombinant hormones can now be produced at low cost and in large amounts. This has led to greater understanding of the pathophysiological factors regulating hematopoiesis. This in turn, hasprompted the search for new therapeutic approaches. EPO might also be used to treat patients with different types of anemia: uremics, newborns, patients with anemia from cancer or myeloproliferative disease, thalassemia, bone marrow transplants, chronic infectious diseases. Besides erythroid cells, EPO affects other blood cell lines, such as myeloid cells, lymphocytes and megakaryocytes. It can also enhance polymorphonuclear cell phagocytosis and reduce macrophage activation, thus modulating the inflammatory process. Hematopoietic and endothelial cells probably have the same origin, and the discovery of eyrthropoietin receptors also on mesangial, myocardial and smooth muscle cells has prompted research into the non-erythropoietic function of the hormone. EPO has an important, direct, hemodynamic and vasoactive effect, which does not depend only on an increase in hematocrit and viscosity. Moreover, EPO and its receptors have been found in the brain, suggesting a role in preventing neuronal death. Finally, the recently discovered interaction between EPO and vascular endothelial growth factor (VEGF), and the ability of EPO to stimulate endothelial cell mitosis and motility may be of importance in neovascularization and wound healing.

  17. Activation of erythropoietin receptor in the absence of hormone by a peptide that binds to a domain different from the hormone binding site

    PubMed Central

    Naranda, Tatjana; Wong, Kenneth; Kaufman, R. Ilene; Goldstein, Avram; Olsson, Lennart

    1999-01-01

    Applying a homology search method previously described, we identified a sequence in the extracellular dimerization site of the erythropoietin receptor, distant from the hormone binding site. A peptide identical to that sequence was synthesized. Remarkably, it activated receptor signaling in the absence of erythropoietin. Neither the peptide nor the hormone altered the affinity of the other for the receptor; thus, the peptide does not bind to the hormone binding site. The combined activation of signal transduction by hormone and peptide was strongly synergistic. In mice, the peptide acted like the hormone, protecting against the decrease in hematocrit caused by carboplatin. PMID:10377456

  18. Recombinant Human Erythropoietin Therapy for a Jehovah's Witness Child With Severe Anemia due to Hemolytic-Uremic Syndrome.

    PubMed

    Woo, Da Eun; Lee, Jae Min; Kim, Yu Kyung; Park, Yong Hoon

    2016-02-01

    Patients with hemolytic-uremic syndrome (HUS) can rapidly develop profound anemia as the disease progresses, as a consequence of red blood cell (RBC) hemolysis and inadequate erythropoietin synthesis. Therefore, RBC transfusion should be considered in HUS patients with severe anemia to avoid cardiac or pulmonary complications. Most patients who are Jehovah's Witnesses refuse blood transfusion, even in the face of life-threatening medical conditions due to their religious convictions. These patients require management alternatives to blood transfusions. Erythropoietin is a glycopeptide that enhances endogenous erythropoiesis in the bone marrow. With the availability of recombinant human erythropoietin (rHuEPO), several authors have reported its successful use in patients refusing blood transfusion. However, the optimal dose and duration of treatment with rHuEPO are not established. We report a case of a 2-year-old boy with diarrhea-associated HUS whose family members are Jehovah's Witnesses. He had severe anemia with acute kidney injury. His lowest hemoglobin level was 3.6 g/dL, but his parents refused treatment with packed RBC transfusion due to their religious beliefs. Therefore, we treated him with high-dose rHuEPO (300 IU/kg/day) as well as folic acid, vitamin B12, and intravenous iron. The hemoglobin level increased steadily to 7.4 g/dL after 10 days of treatment and his renal function improved without any complications. To our knowledge, this is the first case of successful rHuEPO treatment in a Jehovah's Witness child with severe anemia due to HUS. PMID:26958070

  19. Role of recombinant human erythropoietin loading chitosan-tripolyphosphate nanoparticles in busulfan-induced genotoxicity: Analysis of DNA fragmentation via comet assay in cultured HepG2 cells.

    PubMed

    Ghassemi-Barghi, Nasrin; Varshosaz, Jaleh; Etebari, Mahmoud; Jafarian Dehkordi, Abbas

    2016-10-01

    Busulfan is one of the most effective chemotherapeutic agents used for the treatment of chronic myeloid leukemia. Busulfan is involved in secondary malignancy due to its genotoxic potential in normal tissues. As an alkylating agent busulfan can cause DNA damage by cross-linking DNAs and DNA and proteins, induces senescence in normal cells via transient depletion of intracellular glutathione (GSH) and subsequently by a continuous increase in reactive oxygen species (ROS) production. Erythropoietin, a glycoprotein widely used against drug induced anemia in cancerous patients and regulates hematopoiesis, has been shown to exert an important cyto-protective effect in many tissues. Recombinant human erythropoietin has been demonstrated to directly limit cell injury and ROS generation during oxidative stress. Furthermore, rhEPO decreased levels of pro-apoptotic factor (Bax) and also increased expression of the anti-apoptotic factor Bcl2. According to EPO's short half-life and requirements for the frequently administration, finding the new strategies to attenuate its side effects is important. The aim of this study was to explore whether rhEPO loading chitosan-tripolyphosphate nanoparticles protects against busulfan-induced genotoxicity in HepG2 cells. For this purpose cells were incubated with busulfan alone, regular rhEPO alone and regular rhEPO and CS-TPP-EPO nanoparticles along with busulfan in pre and co-treatment condition. Our results showed that busulfan induced a noticeable genotoxic effects in HepG2 cells (p<0.0001). Both regular rhEPO and CS-TPP-EPO nanoparticles reduced the effects of busulfan significantly (p<0.0001) by reduction of the level of DNA damage via blocking ROS generation, and enhancement intracellular glutathione levels. CS-TPP-EPO nanoparticles were more effective than regular rhEPO in both pre and co-treatment conditions. In conclusion, our results show that administration of rhEPO and CS-TPP-EPO nanoparticles especially in the pre

  20. Serum erythropoietin concentrations in symptomatic infants during the anaemia of prematurity.

    PubMed

    Meyer, J; Sive, A; Jacobs, P

    1992-07-01

    A comparison was carried out between a series of neonates who weighed less than 1500 g at birth and received red cell transfusions for symptomatic anaemia of prematurity (group 1, n = 14) and controls of similar gestational age and weight, who remained well and were not transfused during their nursery stay (group 2, n = 10). Mean (SD) haemoglobin concentrations at birth were 163 (12) g/l and 183 (17) g/l (p = 0.004), respectively. Transfusion resulted in significantly better weight gain in six infants who had been growing poorly:mean (SE) 8.8 (2.8) g/day improved to 23.3 (2.1) g/day (p less than 0.002). Geometric mean (SD) serum immunoreactive erythropoietin (SiEp) concentrations (17.7 (1.3) U/l) for the whole group of infants were similar to those of normal adults (17.4 (4.7) U/l) despite considerably reduced haemoglobin values. There was a significant inverse correlation between haemoglobin and log SiEp concentrations in the infants requiring transfusion (r = -0.43; p less than 0.01), but this was not apparent in the untransfused babies. Moreover, at haemoglobin concentrations below 120 g/l the mean (SE) SiEp concentration of 20 (1.08) U/l in group 1 was significantly higher than in group 2 (14 (1.06) U/l; p = 0.002). These data suggest that an increased concentration of SiEp early in the course of the anaemia of prematurity helps to identify those infants who would benefit from red cell transfusions, but that clinical criteria, although ill defined, do so equally well. PMID:1519982

  1. Cost-effectiveness of recombinant human erythropoietin in the prevention of chemotherapy-induced anaemia.

    PubMed Central

    Barosi, G.; Marchetti, M.; Liberato, N. L.

    1998-01-01

    Recombinant human erythropoietin (rHuEPO) has been advocated for the treatment of anaemia in patients submitted to cancer chemotherapy. We used decision analysis to compare the cost-effectiveness of rHuEPO supplemented with red blood cell (RBC) transfusions with conventional treatment with RBC transfusions alone. At baseline, we analysed the use of rHuEPO as secondary prophylaxis according to effectiveness estimates from a community-based oncology study. In order to reduce the probability of transfusions from 21.9% to 10.4%, and the number of RBC units per patient per month from 0.55 to 0.29, 150 units kg(-1) s.c. rHuEPO three times per week for 4 months resulted in an incremental cost of $189,652 per quality-adjusted life year (QALY). In patients treated with cisplatin-containing chemotherapy, rHuEPO added $190,142 per QALY. In a hypothetical scenario of a transfusion pattern that maintained the same haemoglobin level of rHuEPO-responsive patients, the marginal cost of rHuEPO was always greater than $100,000 per QALY. Results were stable with regard to variations in the probability of blood-borne infections, quality of life of responding patients and cancer-related mortality. The additional cost could be lowered to less than $100,000 per QALY by saving 4.5 RBC units over 4 months for any patient treated. In conclusion, according to current use, rHuEPO is not cost-effective in the treatment of chemotherapy-induced anaemia. More tailored utilization of the drug and better consideration of predictive response indicators may lead to an effective, blood-sparing alternative. PMID:9743301

  2. Diurnal changes of arterial oxygen saturation and erythropoietin concentration in male and female highlanders.

    PubMed

    Cristancho, Edgar; Riveros, Alain; Sánchez, Armando; Peñuela, Oscar; Böning, Dieter

    2016-09-01

    In Caucasians and Native Americans living at altitude, hemoglobin mass is increased in spite of erythropoietin concentrations ([Epo]) not markedly differing from sea level values. We hypothesized that a nocturnal decrease of arterial oxygen saturation (SaO2) causes a temporary rise of [Epo] not detected by morning measurements. SaO2 (continuous, finger oximeter) and [Epo] (ELISA, every 4 h) were determined in young highlanders (altitude 2600 m) during 24 h of usual daily activity. In Series I (six male, nine female students), SaO2 fell during the night with the nadir occurring between 01:00 and 03:00; daily means (range 92.4-95.2%) were higher in females (+1.7%, P < 0.01). [Epo] showed opposite changes with zenith occurring at 04:00 without a sex difference. Mean daily values (22.9 ± 10.7SD U/L) were higher than values obtained at 08:00 (17.2 ± 9.5 U/L, P < 0.05). In Series II (seven females), only SaO2 was measured. During follicular and luteal phases, SaO2 variation was similar to Series I, but the rhythm was disturbed during menstruation. While daily [Epo] variations at sea level are not homogeneous, there is a diurnal variation at altitude following changes in SaO2 Larger hypoventilation-dependent decreases of alveolar PO2 decreases during the night probably cause a stronger reduction of SaO2 in highlanders compared to lowlanders. This variation might be enlarged by a diurnal fluctuation of Hb concentration. In spite of a lower [Hb], the higher SaO2 in women compared to men led to a similar arterial oxygen content, likely explaining the absence of differences in [Epo] between sexes. PMID:27597764

  3. Neural stem cell transplantation combined with erythropoietin for the treatment of spinal cord injury in rats

    PubMed Central

    Zhao, Yan; Zuo, Yuan; Jiang, Jianming; Yan, Huibo; Wang, Xiliang; Huo, Hunjun; Xiao, Yulong

    2016-01-01

    Spinal cord injury (SCI) comprises nerve and motor function disorders that may be caused by a variety of damaging factors and is challenging to treat. The aim of the present study was to investigate the regenerative effects of neural stem cell (NSC) transplantation combined with intraperitoneal injection of erythropoietin (EPO) on cross-sectional SCI in rats. A model of SCI was induced in 40 adult Wistar rats via the complete transection of the 10th thoracic vertebra (T10). The rats were allocated at random into 4 groups: Control, NSC, EPO and NSC + EPO groups (n=10 per group). Morphological alterations associated with axonal regeneration were detected using neurofilament (NF)-200 immunohistochemistry and immunofluorescence staining after 8 weeks. Basso, Beattie and Bresnahan (BBB) scoring was used to evaluate the recovery of hindlimb function. A total of 5 rats died following surgery, including 2 control rats and 1 rat each in the EPO, NSC and NSC + EPO groups. NSCs labeled with bromodeoxyuridine were observed to have survived and migrated in the spinal cord tissue after 8 weeks. Significant histomorphological differences were observed in the NSC and NSC + EPO groups compared with the EPO and control groups. Furthermore, the rats of the NSC + EPO group exhibited significantly enhanced axonal regeneration in the SCI area compared with the NSC group rats. The rats of the NSC and NSC + EPO groups exhibited significantly improved BBB scores compared with the EPO and control group rats at 7 days after treatment (P<0.05). In addition, the BBB scores of the NSC + EPO group were significantly improved compared with those of the three other groups at 7 days after surgery (P<0.05). Therefore, the results of the present study suggest that NSC transplantation combined with intraperitoneal injection of EPO may benefit the survival and regeneration of injured axons, and accelerate the repair of injured spinal cord tissue, thus facilitating the functional recovery of hindlimb

  4. Cerebral tissue repair and atrophy after embolic stroke in rat: an MRI study of Erythropoietin therapy

    PubMed Central

    Ding, Guangliang; Jiang, Quan; Li, Lian; Zhang, Li; Wang, Ying; Zhang, Zheng Gang; Lu, Mei; Panda, Swayamprava; Li, Qingjiang; Ewing, James R.; Chopp, Michael

    2010-01-01

    Using magnetic resonance imaging (MRI) protocols of T2-, T2*-, diffusion- and susceptibility-weighted imaging (T2WI, T2*WI, DWI and SWI) with a 7T system, we tested the hypothesis that treatment of embolic stroke with Erythropoietin (EPO) initiated at 24 hours and administered daily for 7 days after stroke onset has benefit on 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 group (n=11). The treated group was given EPO intraperitoneally at a dose of 5,000 IU/kg daily for 7 days starting 24h after MCAO. Controls were given an equal volume of saline. MRI was performed at 24h and then weekly for 6 weeks. MRI and histological measurements were compared between groups. Serial T2WI images 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 24h post stroke promotes angiogenesis and axonal remodeling in the ischemic boundary, which may potentially reduce atrophy of the ipsilateral hemisphere. PMID:20722071

  5. Erythropoietin Modulates Cerebral and Serum Degradation Products from Excess Calpain Activation following Prenatal Hypoxia-Ischemia.

    PubMed

    Jantzie, Lauren L; Winer, Jesse L; Corbett, Christopher J; Robinson, Shenandoah

    2016-01-01

    Preterm infants suffer central nervous system (CNS) injury from hypoxia-ischemia and inflammation - termed encephalopathy of prematurity. Mature CNS injury activates caspase and calpain proteases. Erythropoietin (EPO) limits apoptosis mediated by activated caspases, but its role in modulating calpain activation has not yet been investigated extensively following injury to the developing CNS. We hypothesized that excess calpain activation degrades developmentally regulated molecules essential for CNS circuit formation, myelination and axon integrity, including neuronal potassium-chloride co-transporter (KCC2), myelin basic protein (MBP) and phosphorylated neurofilament (pNF), respectively. Further, we predicted that post-injury EPO treatment could mitigate CNS calpain-mediated degradation. Using prenatal transient systemic hypoxia-ischemia (TSHI) in rats to mimic CNS injury from extreme preterm birth, and postnatal EPO treatment with a clinically relevant dosing regimen, we found sustained postnatal excess cortical calpain activation following prenatal TSHI, as shown by the cleavage of alpha II-spectrin (αII-spectrin) into 145-kDa αII-spectrin degradation products (αII-SDPs) and p35 into p25. Postnatal expression of the endogenous calpain inhibitor calpastatin was also reduced following prenatal TSHI. Calpain substrate expression following TSHI, including cortical KCC2, MBP and NF, was modulated by postnatal EPO treatment. Calpain activation was reflected in serum levels of αII-SDPs and KCC2 fragments, and notably, EPO treatment also modulated KCC2 fragment levels. Together, these data indicate that excess calpain activity contributes to the pathogenesis of encephalopathy of prematurity. Serum biomarkers of calpain activation may detect ongoing cerebral injury and responsiveness to EPO or similar neuroprotective strategies. PMID:26551007

  6. Soluble expression and partial purification of recombinant human erythropoietin from E. coli.

    PubMed

    Jeong, Taeck-Hyun; Son, Young-Jin; Ryu, Han-Bong; Koo, Bon-Kyung; Jeong, Seung-Mi; Hoang, Phuong; Do, Bich Hang; Song, Jung-A; Chong, Seon-Ha; Robinson, Robert Charles; Choe, Han

    2014-03-01

    Human erythropoietin (hEpo) is an essential regulator of erythrocyte production that induces the division and differentiation of erythroid progenitor cells in the bone marrow into mature erythrocytes. It is widely used for the treatment of anemia resulting from chronic kidney disease, chemotherapy, and cancer-related therapies. Active hEpo, and hEpo analogs, have been purified primarily from mammalian cells, which has several disadvantages, including low yields and high production costs. Although an Escherichia coli (E. coli) expression system may provide economic production of therapeutic proteins, it has not been used for the production of recombinant hEpo (rhEpo) because it aggregates in inclusion bodies in the E. coli cytoplasm and is not modified post-translationally. We investigated the soluble overexpression of active rhEpo with various protein tags in E. coli, and found out that several tags increased the solubility of rhEpo. Among them maltose binding protein (MBP)-tagged rhEpo was purified using affinity and gel filtration columns. Non-denaturing electrophoresis and MALDI-TOF MS analysis demonstrated that the purified rhEpo had two intra-disulfide bonds identical to those of the native hEpo. An in vitro proliferation assay showed that rhEpo purified from E. coli had similar biological activity as rhEpo derived from CHO cells. Therefore, we report for the first time that active rhEpo was overexpressed as a soluble form in the cytoplasm of E. coli and purified it in simple purification steps. We hope that our results offer opportunities for progress in rhEpo therapeutics. PMID:24412408

  7. Clinical significance of erythropoietin receptor expression in oral squamous cell carcinoma

    PubMed Central

    2012-01-01

    Background Hypoxic tumors are refractory to radiation and chemotherapy. High expression of biomarkers related to hypoxia in head and neck cancer is associated with a poorer prognosis. The present study aimed to evaluate the clinicopathological significance of erythropoietin receptor (EPOR) expression in oral squamous cell carcinoma (OSCC). Methods The study included 256 patients who underwent primary surgical resection between October 1996 and August 2005 for treatment of OSCC without previous radiotherapy and/or chemotherapy. Clinicopathological information including gender, age, T classification, N classification, and TNM stage was obtained from clinical records and pathology reports. The mRNA and protein expression levels of EPOR in OSCC specimens were evaluated by Q-RT-PCR, Western blotting and immunohistochemistry assays. Results We found that EPOR were overexpressed in OSCC tissues. The study included 17 women and 239 men with an average age of 50.9 years (range, 26–87 years). The mean follow-up period was 67 months (range, 2–171 months). High EPOR expression was significantly correlated with advanced T classification (p < 0.001), advanced TNM stage (p < 0.001), and positive N classification (p = 0.001). Furthermore, the univariate analysis revealed that patients with high tumor EPOR expression had a lower 5-year overall survival rate (p = 0.0011) and 5-year disease-specific survival rate (p = 0.0017) than patients who had low tumor levels of EPOR. However, the multivariate analysis using Cox’s regression model revealed that only the T and N classifications were independent prognostic factors for the 5-year overall survival and 5-year disease-specific survival rates. Conclusions High EPOR expression in OSCC is associated with an aggressive tumor behavior and poorer prognosis in the univariate analysis among patients with OSCC. Thus, EPOR expression may serve as a treatment target for OSCC in the future. PMID:22639817

  8. The Neuroprotective Mechanism of Erythropoietin-TAT Fusion Protein Against Neurodegeneration from Ischemic Brain Injury.

    PubMed

    Liu, P; Liu, X; Akf Liou, Emptyyn Y; Xing, J; Jing, Z; Ji, X; Liu, X; Zhao, H; Yan, F; Chen, J; Cao, G; Luo, Y

    2013-08-27

    Aims: To compare the neuroprotection of erythropoietin (EPO) and EPO fusion protein containing transduction domain derived from HIV TAT (EPO-TAT) against ischemic brain injury, inclusive of the side effect, and explore the mechanism underlying the role of EPO-TAT in a transient focal cerebral ischemia model in rats. Methods: Transient focal ischemia was induced by middle cerebral artery occlusion (MCAO) in rats. Rats were treated, respectively, with following regimens: saline, 1000 U/kg EPO, 5000 U/kg EPO, 1000 U/kg EPO-TAT, 1000 U/kg EPO-TAT + 5 µl of 10 mM LY294002 (or/plus 5 µl of 5 mM PD98059). Neurological deficit scores, infarct volume, and hematologic side effect were assessed at 72 hours after MCAO. Apoptotic cells were determined with TUNEL staining. The expression and localization of phosphorylated AKT (pAKT) and phosphorylated ERK (pERK) were detected with Western blot, immunohistochemistry, and immunofluorescence, respectively. Results: 1000 U/kg EPO-TAT exhibited a comparable neuroprotection to 5000 U/kg EPO, as evidenced by a comparable attenuation in neurological deficit, infarct volume, and number of apoptotic cells in the rat ischemic cortex after MCAO. The pAKT and pERK levels were significantly elevated solely in neurons of rodents receiving EPO or EPO-TAT treatments, suggesting the concurrent activation of these two pathways. Specific inhibition of either AKT or ERK pathway partially abolished EPO-TAT protection, but exhibited no influence on the activation status of its counterpart, suggesting no cross-modulation between these two protective pathways. Conclusion: Our study indicates that EPO-TAT at 1000 U/kg displays neuroprotection with no detectable side effects. The mechanism for neuroprotection may be attributable to the simultaneous activation of the AKT and ERK pathways, which preserve neuronal cell viability and attenuate behavioral deficits. PMID:24040790

  9. THE COMBINATION OF α-LIPOIC ACID INTAKE WITH ECCENTRIC EXERCISE MODULATES ERYTHROPOIETIN RELEASE.

    PubMed

    Morawin, B; Turowski, D; Naczk, M; Siatkowski, I; Zembron-Lacny, A

    2014-08-01

    The generation of reactive nitrogen/oxygen species (RN/OS) represents an important mechanism in erythropoietin (EPO) expression and skeletal muscle adaptation to physical and metabolic stress. RN/OS generation can be modulated by intense exercise and nutrition supplements such as α-lipoic acid, which demonstrates both anti- and pro-oxidative action. The study was designed to show the changes in the haematological response through the combination of α-lipoic acid intake with running eccentric exercise. Sixteen healthy young males participated in the randomised and placebo-controlled study. The exercise trial involved a 90-min run followed by a 15-min eccentric phase at 65% VO2max (-10% gradient). It significantly increased serum concentrations of nitric oxide (NO), hydrogen peroxide (H2O2) and pro-oxidative products such as 8-isoprostanes (8-iso), lipid peroxides (LPO) and protein carbonyls (PC). α-Lipoic acid intake (Thiogamma: 1200 mg daily for 10 days prior to exercise) resulted in a 2-fold elevation of serum H2O2 concentration before exercise, but it prevented the generation of NO, 8-iso, LPO and PC at 20 min, 24 h, and 48 h after exercise. α-Lipoic acid also elevated serum EPO level, which highly correlated with NO/H2O2 ratio (r = 0.718, P < 0.01). Serum total creatine kinase (CK) activity, as a marker of muscle damage, reached a peak at 24 h after exercise (placebo 732 ± 207 IU · L(-1), α-lipoic acid 481 ± 103 IU · L(-1)), and correlated with EPO (r = 0.478, P < 0.01) in the α-lipoic acid group. In conclusion, the intake of high α-lipoic acid modulates RN/OS generation, enhances EPO release and reduces muscle damage after running eccentric exercise.

  10. Recognition and characterization of Erythropoietin binding-proteins in the brain of mice

    PubMed Central

    Kowsari, Reza; Yazdian-Robati, Rezvan; Razavi, Bibi Marjan; Pourtaji, Atena; Ghorbani, Maryam; Moghadam-Omranipour, Hediye; Hosseinzadeh, Hossein; Lari, Parisa; Abnous, Khalil

    2016-01-01

    Objective(s): Erythropoietin (EPO), is a 34KDa glycoprotein hormone, which belongs to type 1 cytokine superfamily. EPO involves in erythrocyte maturation through inhibition of apoptosis in erythroid cells. Besides its main function, protective effects of EPO in heart and brain tissues have been reported. EPO has a critical role in development, growth, and homeostasis of brain. Furthermore EPO has great potential in the recovery of different brain diseases which are still under studying. In this research, EPO binding pattern to brain proteins in animal model was studied. Materials and Methods: EPO antibody was covalently crosslinked to protein A/G agarose. in order to interact between EPO and its target in brain, about 5µg EPO added to brain homogenates(500ul of 1 mg/ml) and incubate at 4ο C for 30 min. brain tissue lysate were added to agarose beads, After isolation of target proteins(EPO - protein) both one and two-dimensional gel electrophoresis were performed. Proteins were identified utilizing MALDI-TOF/TOF and MASCOT software. Results: This research showed that EPO could physically interact with eightproteins including Tubulin beta, Actin cytoplasmic 2, T-complex protein 1, TPR and ankyrin repeat-containing protein 1, Centromere-associated protein E, Kinesin-like protein KIF7, Growth arrest-specific protein 2 and Pleckstrin homology-like domain family B member 2. Conclusion: Since EPO is a promising therapeutic drug for the treatment of neurological diseases, identified proteins may help us to have a better understanding about the mechanism of protective effects of EPO in the brain. Our data needs to be validated by complementary bioassays. PMID:27803781

  11. Ventilation, autonomic function, sleep and erythropoietin. Chronic mountain sickness of Andean natives.

    PubMed

    Bernardi, Luciano; Roach, Robert C; Keyl, Cornelius; Spicuzza, Lucia; Passino, Claudio; Bonfichi, Maurizio; Gamboa, Alfredo; Gamboa, Jorge; Malcovati, Luca; Schneider, Annette; Casiraghi, Nadia; Mori, Antonio; Leon-Velarde, Fabiola

    2003-01-01

    Polycythemia is one of the key factors involved in the chronic mountain sickness syndrome, a condition frequent in Andean natives but whose causes still remain unclear. In theory, polycythemia may be secondary to abnormalities in ventilation, occurring during day or night (e.g. due to sleep abnormalities) stimulating excessive erythropoietin (Epo) production, or else it may result from either autogenous production, or from co-factors like cobalt. To assess the importance of these points, we studied subjects with or without polycythemia, born and living in Cerro de Pasco (Peru, 4330m asl, CP) and evaluated the relationship between Epo and respiratory variables both in CP and sea level. We also assessed the relationship between sleep abnormalities and the circadian rhythm of Epo. Polycythemic subjects showed higher Epo in all conditions, lower SaO2 and hypoxic ventilatory response, higher physiological dead space and higher CO2, suggesting ventilatory inefficiency. Epo levels could be highly modified by the level of oxygenation, and were related to similar directional changes in SaO2. Cobalt levels were normal in all subjects and correlated poorly with hematologic variables. The diurnal variations in Epo were grossly abnormal in polycythemic subjects, with complete loss of the circadian rhythm. These abnormalities correlated with the levels of hypoxemia during the night, but not with sleep abnormalities, which were only minor even in polycythemic subjects. The increased Epo production is mainly related to a greater ventilatory inefficiency, and not to altered sensitivity to hypoxia, cobalt or sleep abnormalities. Improving oxygenation can represent a possible therapeutic option for this syndrome.

  12. Decreased plasma soluble erythropoietin receptor in high-altitude excessive erythrocytosis and Chronic Mountain Sickness

    PubMed Central

    Macarlupú, José Luis; Anza-Ramírez, Cecilia; Corrales-Melgar, Daniela; Vizcardo-Galindo, Gustavo; Corante, Noemí; León-Velarde, Fabiola

    2014-01-01

    Excessive erythrocytosis (EE) is the hallmark of chronic mountain sickness (CMS), a prevalent syndrome in high-altitude Andean populations. Although hypoxemia represents its underlying stimulus, why some individuals develop EE despite having altitude-normal blood erythropoietin (Epo) concentration is still unclear. A soluble form of the Epo receptor (sEpoR) has been identified in human blood and competes directly for Epo with its membrane counterpart (mEpoR). Thus, reduced levels of circulating sEpoR could lead to higher Epo availability and ultimately to EE. We characterized the relationship between Epo and sEpoR, with hematocrit and hemoglobin concentration in healthy highlanders and CMS patients at 4,340 m in Cerro de Pasco, Peru. Our results show that EE patients show decreased plasma sEpoR levels and can be subdivided into two subgroups of normal and high plasma Epo concentration for the altitude of residence, with hemoglobin concentration rising exponentially with an increasing Epo-to-sEpoR ratio (Epo/sEpoR). Also, we showed that the latter varies as an inverse exponential function of arterial pulse O2 saturation. Our findings suggests that EE is strongly associated with higher Epo/sEpoR values, leading to elevated plasma Epo availability to bind mEpoR, and thereby a stronger stimulus for augmented erythropoiesis. Differences in the altitude normal and high Epo CMS patients with a progressively higher Epo/sEpoR supports the hypothesis of the existence of two genetically different subgroups suffering from EE and possibly different degrees of adaptation to chronic high-altitude hypoxia. PMID:25324511

  13. Decreased plasma soluble erythropoietin receptor in high-altitude excessive erythrocytosis and Chronic Mountain Sickness.

    PubMed

    Villafuerte, Francisco C; Macarlupú, José Luis; Anza-Ramírez, Cecilia; Corrales-Melgar, Daniela; Vizcardo-Galindo, Gustavo; Corante, Noemí; León-Velarde, Fabiola

    2014-12-01

    Excessive erythrocytosis (EE) is the hallmark of chronic mountain sickness (CMS), a prevalent syndrome in high-altitude Andean populations. Although hypoxemia represents its underlying stimulus, why some individuals develop EE despite having altitude-normal blood erythropoietin (Epo) concentration is still unclear. A soluble form of the Epo receptor (sEpoR) has been identified in human blood and competes directly for Epo with its membrane counterpart (mEpoR). Thus, reduced levels of circulating sEpoR could lead to higher Epo availability and ultimately to EE. We characterized the relationship between Epo and sEpoR, with hematocrit and hemoglobin concentration in healthy highlanders and CMS patients at 4,340 m in Cerro de Pasco, Peru. Our results show that EE patients show decreased plasma sEpoR levels and can be subdivided into two subgroups of normal and high plasma Epo concentration for the altitude of residence, with hemoglobin concentration rising exponentially with an increasing Epo-to-sEpoR ratio (Epo/sEpoR). Also, we showed that the latter varies as an inverse exponential function of arterial pulse O2 saturation. Our findings suggests that EE is strongly associated with higher Epo/sEpoR values, leading to elevated plasma Epo availability to bind mEpoR, and thereby a stronger stimulus for augmented erythropoiesis. Differences in the altitude normal and high Epo CMS patients with a progressively higher Epo/sEpoR supports the hypothesis of the existence of two genetically different subgroups suffering from EE and possibly different degrees of adaptation to chronic high-altitude hypoxia.

  14. Erythropoietin pharmacokinetic/pharmacodynamic analysis suggests higher doses in treating neonatal anemia

    PubMed Central

    Neelakantan, Srividya; Widness, John A.; Schmidt, Robert L.; Veng-Pedersen, Peter

    2010-01-01

    Background The establishment of effective treatment of neonatal anemia using recombinant human erythropoietin (r-HuEPO) requires a thorough understanding of the physiology and mechanism of EPO’s pharmacologic effect. The purpose of the present preclinical study in sheep was to elucidate the stimulatory effect of EPO on erythroid progenitors and their differentiation into reticulocytes useful in predicting optimal r-HuEPO dosing. Methods Five young adult sheep each underwent two phlebotomies spaced 4–6 weeks apart in which their hemoglobin levels were reduced from 12 g/dL to 3–4 g/dL. Endogenous EPO levels and reticulocyte counts produced in response to anemia were sampled throughout the study and analyzed using a pharmacokinetic/pharmacodynamic (PK/PD) model. Results The phlebotomy-induced drop in hemoglobin resulted in a increase in EPO levels, which reached a maximum of 764 ± 55 mU/mL (mean ± %CV) in 0.5–2.6 days. The reticulocyte counts increased from baseline values of 76.9 × 103 ± 67/μL to 619 × 103 ± 30/μL in 8 days. The PK/PD analysis indicated an increased maturation time for the reticulocytes (4.88 ± 35 days) and demonstrated that the Emax model for EPO’s activation of the progenitors did not show significant effect saturation at the endogenous EPO levels reached. Conclusions In extrapolating from the animal pilot experiment, the present study provides a case for the use of higher r-HuEPO doses in human studies to determine if higher doses are more effective in treatment of neonatal anemia to reduce, and in some less severe cases, eliminate, the need for blood transfusions. PMID:19371274

  15. The Greatly Misunderstood Erythropoietin Resistance Index and the Case for a New Responsiveness Measure

    PubMed Central

    Chait, Y; Kalim, S; Horowitz, J; Hollot, CH; Ankers, ED; Germain, MJ; Thadhani, RI

    2016-01-01

    Background The optimal use of erythropoiesis stimulating agents (ESAs) to treat anemia in end stage renal disease (ESRD) remains controversial due to reported associations with adverse events. In analyzing these associations, studies often utilize ESA resistance indices (ERIs), to characterize a patient’s response to ESA. In this study, we examine whether ERI is an adequate measure of ESA resistance. Methods We used retrospective data from a non-concurrent cohort study of incident hemodialysis patients in the United States (n=9386). ERI is defined as average weekly erythropoietin (EPO) dose per kg body weight (wt) per average hemoglobin (Hgb), over a 3-month period (ERI = (EPO/wt)/Hgb). Linear regression was used to demonstrate the relationship between ERI and weight-adjusted EPO. The coefficient of variation (CV) was used to compare the variability of Hgb with that of weight-adjusted EPO in order to explain this relationship. This analysis was done for each quarter during the first year of dialysis. Results ERI is strongly linearly related with weight-adjusted EPO dose in each of the 4 quarters by the equation ERI = 0.0899*(EPO/wt) (range of R2 = 0.97–0.98) and weakly linearly related to 1/Hgb (range of R2 = 0.06–0.16). These correlations hold independent of age, sex, hgb level, ERI level, and epo-naïve stratifications. Conclusions ERI is strongly linearly related to weight-adjusted (and non-weight-adjusted) EPO dose by a “universal”, not patient-specific formula, and thus is a surrogate of EPO dose. Therefore, associations between ERI and clinical outcomes are associations between a confounded EPO dose and those outcomes. PMID:26843352

  16. Erythropoietin Synthesis in Renal Myofibroblasts Is Restored by Activation of Hypoxia Signaling.

    PubMed

    Souma, Tomokazu; Nezu, Masahiro; Nakano, Daisuke; Yamazaki, Shun; Hirano, Ikuo; Sekine, Hiroki; Dan, Takashi; Takeda, Kotaro; Fong, Guo-Hua; Nishiyama, Akira; Ito, Sadayoshi; Miyata, Toshio; Yamamoto, Masayuki; Suzuki, Norio

    2016-02-01

    Erythropoietin (Epo) is produced by renal Epo-producing cells (REPs) in a hypoxia-inducible manner. The conversion of REPs into myofibroblasts and coincident loss of Epo-producing ability are the major cause of renal fibrosis and anemia. However, the hypoxic response of these transformed myofibroblasts remains unclear. Here, we used complementary in vivo transgenic and live imaging approaches to better understand the importance of hypoxia signaling in Epo production. Live imaging of REPs in transgenic mice expressing green fluorescent protein from a modified Epo-gene locus revealed that healthy REPs tightly associated with endothelium by wrapping processes around capillaries. However, this association was hampered in states of renal injury-induced inflammation previously shown to correlate with the transition to myofibroblast-transformed renal Epo-producing cells (MF-REPs). Furthermore, activation of hypoxia-inducible factors (HIFs) by genetic inactivation of HIF-prolyl hydroxylases (PHD1, PHD2, and PHD3) selectively in Epo-producing cells reactivated Epo production in MF-REPs. Loss of PHD2 in REPs restored Epo-gene expression in injured kidneys but caused polycythemia. Notably, combined deletions of PHD1 and PHD3 prevented loss of Epo expression without provoking polycythemia. Mice with PHD-deficient REPs also showed resistance to LPS-induced Epo repression in kidneys, suggesting that augmented HIF signaling counterbalances inflammatory stimuli in regulation of Epo production. Thus, augmentation of HIF signaling may be an attractive therapeutic strategy for treating renal anemia by reactivating Epo synthesis in MF-REPs.

  17. Establishment of the first WHO Erythropoietin antibody reference panel: Report of an international collaborative study.

    PubMed

    Wadhwa, Meenu; Mytych, Daniel T; Bird, Chris; Barger, Troy; Dougall, Thomas; Han, Hong; Rigsby, Peter; Kromminga, Arno; Thorpe, Robin

    2016-08-01

    A panel of 9 fully human monoclonal antibodies against human erythropoietin (EPO) with defined characteristics (non-neutralizing, neutralizing, various isotypes, affinities) representative of those evident in antibody-mediated pure red cell aplasia (PRCA) and non-PRCA patients were formulated and lyophilized. The panel was evaluated in a multi-centre international collaborative study comprising eighteen different laboratories using different assay platforms including those in routine use. These included binding assays, some based on use of novel technologies and neutralization assays predominantly employing EPO responsive cell-lines. Results showed that detection and titre varied depending on antibody characteristics and the method used. Only selective assay platforms were capable of detecting the diverse repertoire of EPO antibodies in the panel indicating that some clinically relevant antibodies are likely to be missed in some assays. Importantly, the clinical samples from PRCA patients were distinguished as antibody-positive and the healthy donor serum as antibody negative across all different platforms tested. For neutralization, data was generally consistent across the assays for the different samples regardless of the cell-line and the assay conditions. The heterogeneity in data from the study clearly indicated the need for reference standards for consistency in detecting and measuring EPO antibodies across different assay platforms for monitoring the safety and efficacy of erythropoiesis stimulating agents. Therefore, the WHO ECBS at its meeting in October'15 established the EPO antibody panel, available from NIBSC, to facilitate decision-making on assay selection for testing antibodies against human EPO, for evaluating assay performance of antibody assays for clinical use, for assay validation and for standardization. PMID:27173074

  18. Effects of recombinant human erythropoietin injections on physical self in endurance athletes.

    PubMed

    Ninot, Grégory; Connes, Philippe; Caillaud, Corrine

    2006-04-01

    This study examined the time course of mean self-esteem and physical self scores in three groups: male endurance athletes treated with recombinant human erythropoietin (rHuEPO group, n = 6), a placebo group (n = 5) injected with a sodium chloride solution and a control group who did not receive any injection (n = 6). Each participant completed the Physical Self Inventory twice a day (between 07.00 and 09.00 h and between 19.00 and 21.00 h). Using a 10 cm visual analog scale, the participants assessed global self-esteem, physical self-worth and the sub-domains of physical condition, sport competence, attractive body and physical strength (Fox & Corbin, 1989). This was conducted over three consecutive periods: in the 2 weeks before the course of injections, during the 6 weeks of injections and for 4 weeks after the injections. Aerobic capacity was assessed before and after 4 weeks of treatment. The results showed a significant increase in aerobic physical fitness in the rHuEPO group and a significant increase in perceived physical condition and physical strength scores at the end of treatment. The main psychological result was that endurance athletes were highly sensitive to the effects of rHuEPO on physical fitness. The perception of increased physical condition may lead to a stronger commitment to training. The rHuEPO injections presented a dangerous hedonic effect linked to endurance training. These results confirm the need to tackle rHuEPO abuse at any time during the training season. PMID:16492602

  19. Cellular trafficking and degradation of erythropoietin and novel erythropoiesis stimulating protein (NESP).

    PubMed

    Gross, Alec W; Lodish, Harvey F

    2006-01-27

    Erythropoietin (Epo) is essential for the production of mature red blood cells, and recombinant Epo is commonly used to treat anemia, but how Epo is degraded and cleared from the body is not understood. Glycosylation of Epo is required for its in vivo bioactivity, although not for in vitro receptor binding or stimulation of Epo-dependent cell lines; Epo glycosylation actually reduces the affinity of Epo for the Epo receptor (EpoR). Interestingly, a hyperglycosylated analog of Epo, called novel erythropoiesis-stimulating protein (NESP), has a lower affinity than Epo for the EpoR but has greater in vivo activity and a longer serum half-life than Epo. We hypothesize that a major mechanism for degradation of Epo in the body occurs in cells expressing the Epo receptor, through receptor-mediated endocytosis of Epo followed by degradation in lysosomes, and therefore investigated the trafficking and degradation of Epo and NESP by EpoR-expressing cells. We show that Epo and NESP are degraded only by cultured cells that express the EpoR, and their receptor binding, dissociation, and trafficking properties determine their rates of intracellular degradation. Epo binds surface EpoR faster than NESP (k(on) = 5.0 x 10(8) m(-1) min(-1) versus 1.1 x 10(8) m(-1) min(-1)) but dissociates slower (k(off) = 0.029 min(-1) versus 0.042 min(-1)). Surface-bound Epo and NESP are internalized at the same rate (k(in) = 0.06 min(-1)), and after internalization 60% of each ligand is resecreted intact and 40% degraded. Our kinetic model of Epo and NESP receptor binding, intracellular trafficking, and degradation explains why Epo is degraded faster than NESP at the cellular level.

  20. Effects of erythropoietin on muscle O2 transport during exercise in patients with chronic renal failure.

    PubMed Central

    Marrades, R M; Roca, J; Campistol, J M; Diaz, O; Barberá, J A; Torregrosa, J V; Masclans, J R; Cobos, A; Rodríguez-Roisin, R; Wagner, P D

    1996-01-01

    Erythropoietin (rHuEPO) has proven to be effective in the treatment of anemia of chronic renal failure (CRF). Despite improving the quality of life, peak oxygen uptake after rHuEPO therapy is not improved as much as the increase in hemoglobin concentration ([Hb)] would predict. We hypothesized that this discrepancy is due to failure of O2 transport rates to rise in a manner proportional to [Hb]. To test this, eight patients with CRF undergoing regular hemodialysis were studied pre- and post-rHuEPO ([Hb] = 7.5 +/- 1.0 vs. 12.5 +/- 1.0 g x dl-1) using a standard incremental cycle exercise protocol. A group of 12 healthy sedentary subjects of similar age and anthropometric characteristics served as controls. Arterial and femoral venous blood gas data were obtained and coupled with simultaneous measurements of femoral venous blood flow (Qleg) by thermodilution to obtain O2 delivery and oxygen uptake (VO2). Despite a 68% increase in [Hb], peak VO2 increased by only 33%. This could be explained largely by reduced peak leg blood flow, limiting the gain in O2 delivery to 37%. At peak VO2, after rHuEPO, O2 supply limitation of maximal VO2 was found to occur, permitting the calculation of a value for muscle O2 conductance from capillary to mitochondria (DO2). While DO2 was slightly improved after rHuEPO, it was only 67% of that of sedentary control subjects. This kept maximal oxygen extraction at only 70%. Two important conclusions can be reached from this study. First, the increase in [Hb] produced by rHuEPO is accompanied by a significant reduction in peak blood flow to exercising muscle, which limits the gain in oxygen transport. Second, even after restoration of [Hb], O2 conductance from the muscle capillary to the mitochondria remains considerably below normal. PMID:8621799

  1. Selectively crosslinked hyaluronic acid hydrogels for sustained release formulation of erythropoietin.

    PubMed

    Motokawa, Keiko; Hahn, Sei Kwang; Nakamura, Teruo; Miyamoto, Hajime; Shimoboji, Tsuyoshi

    2006-09-01

    A novel sustained release formulation of erythropoietin (EPO) was developed using hyaluronic acid (HA) hydrogels. For the preparation of HA hydrogels, adipic acid dihydrazide grafted HA (HA-ADH) was synthesized and analyzed with (1)H NMR. The degree of HA-ADH modification was about 69%. EPO was in situ encapsulated into HA-ADH hydrogels through a selective cross-linking reaction of bis(sulfosuccinimidyl) suberate (BS(3)) to hydrazide group (pK(a) = 3.0) of HA-ADH rather than to amine group (pK(a) > 9) of EPO. The denaturation of EPO during HA-ADH hydrogel synthesis was drastically reduced with decreasing pH from 7.4 to 4.8. The specific reactivity of BS(3) to hydrazide at pH = 4.8 might be due to its low pK(a) compared with that of amine. In vitro release of EPO in phosphate buffered saline at 37 degrees C showed that EPO was released rapidly for 2 days and then slowly up to 4 days from HA-ADH hydrogels. When the hydrogels were dried at 37 degrees C for a day, however, longer release of EPO up to 3 weeks could be demonstrated. According to in vivo release test of EPO from HA-ADH hydrogels in SD rats, elevated EPO concentration higher than 0.1 ng/mL could be maintained from 7 days up to 18 days depending on the preparation methods of HA-ADH hydrogels. There was no adverse effect during and after HA-ADH hydrogel implantation. PMID:16721757

  2. Neural stem cell transplantation combined with erythropoietin for the treatment of spinal cord injury in rats

    PubMed Central

    Zhao, Yan; Zuo, Yuan; Jiang, Jianming; Yan, Huibo; Wang, Xiliang; Huo, Hunjun; Xiao, Yulong

    2016-01-01

    Spinal cord injury (SCI) comprises nerve and motor function disorders that may be caused by a variety of damaging factors and is challenging to treat. The aim of the present study was to investigate the regenerative effects of neural stem cell (NSC) transplantation combined with intraperitoneal injection of erythropoietin (EPO) on cross-sectional SCI in rats. A model of SCI was induced in 40 adult Wistar rats via the complete transection of the 10th thoracic vertebra (T10). The rats were allocated at random into 4 groups: Control, NSC, EPO and NSC + EPO groups (n=10 per group). Morphological alterations associated with axonal regeneration were detected using neurofilament (NF)-200 immunohistochemistry and immunofluorescence staining after 8 weeks. Basso, Beattie and Bresnahan (BBB) scoring was used to evaluate the recovery of hindlimb function. A total of 5 rats died following surgery, including 2 control rats and 1 rat each in the EPO, NSC and NSC + EPO groups. NSCs labeled with bromodeoxyuridine were observed to have survived and migrated in the spinal cord tissue after 8 weeks. Significant histomorphological differences were observed in the NSC and NSC + EPO groups compared with the EPO and control groups. Furthermore, the rats of the NSC + EPO group exhibited significantly enhanced axonal regeneration in the SCI area compared with the NSC group rats. The rats of the NSC and NSC + EPO groups exhibited significantly improved BBB scores compared with the EPO and control group rats at 7 days after treatment (P<0.05). In addition, the BBB scores of the NSC + EPO group were significantly improved compared with those of the three other groups at 7 days after surgery (P<0.05). Therefore, the results of the present study suggest that NSC transplantation combined with intraperitoneal injection of EPO may benefit the survival and regeneration of injured axons, and accelerate the repair of injured spinal cord tissue, thus facilitating the functional recovery of hindlimb

  3. Lenalidomide induces lipid raft assembly to enhance erythropoietin receptor signaling in myelodysplastic syndrome progenitors.

    PubMed

    McGraw, Kathy L; Basiorka, Ashley A; Johnson, Joseph O; Clark, Justine; Caceres, Gisela; Padron, Eric; Heaton, Ruth; Ozawa, Yukiyasu; Wei, Sheng; Sokol, Lubomir; List, Alan F

    2014-01-01

    Anemia remains the principal management challenge for patients with lower risk Myelodysplastic Syndromes (MDS). Despite appropriate cytokine production and cellular receptor display, erythropoietin receptor (EpoR) signaling is impaired. We reported that EpoR signaling is dependent upon receptor localization within lipid raft microdomains, and that disruption of raft integrity abolishes signaling capacity. Here, we show that MDS erythroid progenitors display markedly diminished raft assembly and smaller raft aggregates compared to normal controls (p = 0.005, raft number; p = 0.023, raft size). Because lenalidomide triggers raft coalescence in T-lymphocytes promoting immune synapse formation, we assessed effects of lenalidomide on raft assembly in MDS erythroid precursors and UT7 cells. Lenalidomide treatment rapidly induced lipid raft formation accompanied by EpoR recruitment into raft fractions together with STAT5, JAK2, and Lyn kinase. The JAK2 phosphatase, CD45, a key negative regulator of EpoR signaling, was displaced from raft fractions. Lenalidomide treatment prior to Epo stimulation enhanced both JAK2 and STAT5 phosphorylation in UT7 and primary MDS erythroid progenitors, accompanied by increased STAT5 DNA binding in UT7 cells, and increased erythroid colony forming capacity in both UT7 and primary cells. Raft induction was associated with F-actin polymerization, which was blocked by Rho kinase inhibition. These data indicate that deficient raft integrity impairs EpoR signaling, and provides a novel strategy to enhance EpoR signal fidelity in non-del(5q) MDS.

  4. Erythropoietin for the Treatment of Subarachnoid Hemorrhage: A Feasible Ingredient for a Successful Medical Recipe

    PubMed Central

    Grasso, Giovanni; Tomasello, Giovanni; Noto, Marcello; Alafaci, Concetta; Cappello, Francesco

    2015-01-01

    Subarachnoid hemorrhage (SAH) following aneurysm bleeding accounts for 6% to 8% of all cerebrovascular accidents. Although an aneurysm can be effectively managed by surgery or endovascular therapy, delayed cerebral ischemia is diagnosed in a high percentage of patients resulting in significant morbidity and mortality. Cerebral vasospasm occurs in more than half of all patients after aneurysm rupture and is recognized as the leading cause of delayed cerebral ischemia after SAH. Hemodynamic strategies and endovascular procedures may be considered for the treatment of cerebral vasospasm. In recent years, the mechanisms contributing to the development of vasospasm, abnormal reactivity of cerebral arteries and cerebral ischemia following SAH, have been investigated intensively. A number of pathological processes have been identified in the pathogenesis of vasospasm, including endothelial injury, smooth muscle cell contraction from spasmogenic substances produced by the subarachnoid blood clots, changes in vascular responsiveness and inflammatory response of the vascular endothelium. To date, the current therapeutic interventions remain ineffective as they are limited to the manipulation of systemic blood pressure, variation of blood volume and viscosity and control of arterial carbon dioxide tension. In this scenario, the hormone erythropoietin (EPO) has been found to exert neuroprotective action during experimental SAH when its recombinant form (rHuEPO) is administered systemically. However, recent translation of experimental data into clinical trials has suggested an unclear role of recombinant human EPO in the setting of SAH. In this context, the aim of the current review is to present current evidence on the potential role of EPO in cerebrovascular dysfunction following aneurysmal subarachnoid hemorrhage. PMID:26581085

  5. Protective action of erythropoietin on neuronal damage induced by activated microglia.

    PubMed

    Wenker, Shirley D; Chamorro, María E; Vittori, Daniela C; Nesse, Alcira B

    2013-04-01

    Inflammation is a physiological defense response, but may also represent a potential pathological process in neurological diseases. In this regard, microglia have a crucial role in either progression or amelioration of degenerative neuronal damage. Because of the role of hypoxia in pro-inflammatory mechanisms in the nervous system, and the potential anti-inflammatory protective effect of erythropoietin (Epo), we focused our investigation on the role of this factor on activation of microglia and neuroprotection. Activation of microglial cells (EOC-2) was achieved by chemical hypoxia induced by cobalt chloride (CoCl2 ) and characterized by increased levels of nitrite, tumor necrosis factor-α and reactive oxygen species production, as well as up-regulation of inducible nitric oxide synthase expression. Under these conditions, cell proliferation data and proliferating cell nuclear antigen (PCNA) staining demonstrated a mitogenic effect of chemical hypoxia. Even though pre-treatment with Epo did not prevent nitrite production, inducible nitric oxide synthase protein expression or tumor necrosis factor-α secretion, it prevented the oxidative stress induced by CoCl2 as well as cell proliferation. Neuronal cells (SH-SY5Y) cultured in the presence of conditioned medium from activated EOC-2 cells or macrophages (RAW 264.7) developed significant apoptosis, an effect that was abolished by Epo via Epo/Epo receptor activation. The results show that even though Epo did not exert a direct anti-inflammatory effect on microglia activation, it did increase the resistance of neurons to subsequent damage from pro-inflammatory agents. In addition to its anti-apoptotic ability, the Epo antioxidant effect may have an indirect influence on neuronal survival by modulation of the pro-inflammatory environment. PMID:23384249

  6. Endoplasmic reticulum stress signal impairs erythropoietin production: a role for ATF4.

    PubMed

    Chiang, Chih-Kang; Nangaku, Masaomi; Tanaka, Tetsuhiro; Iwawaki, Takao; Inagi, Reiko

    2013-02-15

    Hypoxia upregulates the hypoxia-inducible factor (HIF) pathway and the endoplasmic reticulum (ER) stress signal, unfolded protein response (UPR). The cross talk of both signals affects the pathogenic alteration by hypoxia. Here we showed that ER stress induced by tunicamycin or thapsigargin suppressed inducible (CoCl(2) or hypoxia) transcription of erythropoietin (EPO), a representative HIF target gene, in HepG2. This suppression was inversely correlated with UPR activation, as estimated by expression of the UPR regulator glucose-regulated protein 78, and restored by an ER stress inhibitor, salubrinal, in association with normalization of the UPR state. Importantly, the decreased EPO expression was also observed in HepG2 overexpressing UPR activating transcription factor (ATF)4. Overexpression of mutated ATF4 that lacks the transcriptional activity did not alter EPO transcriptional regulation. Transcriptional activity of the EPO 3'-enhancer, which is mainly regulated by HIF, was abolished by both ER stressors and ATF4 overexpression, while nuclear HIF accumulation or expression of other HIF target genes was not suppressed by ER stress. Chromatin immunoprecipitation analysis identified a novel ATF4 binding site (TGACCTCT) within the EPO 3'-enhancer region, suggesting a distinct role for ATF4 in UPR-dependent suppression of the enhancer. Induction of ER stress in rat liver and kidney by tunicamycin decreased the hepatic and renal mRNA and plasma level of EPO. Collectively, ER stress selectively impairs the transcriptional activity of EPO but not of other HIF target genes. This effect is mediated by suppression of EPO 3'-enhancer activity via ATF4 without any direct effect on HIF, indicating that UPR contributes to oxygen-sensing regulation of EPO. PMID:23242184

  7. Erythropoietin Synthesis in Renal Myofibroblasts Is Restored by Activation of Hypoxia Signaling.

    PubMed

    Souma, Tomokazu; Nezu, Masahiro; Nakano, Daisuke; Yamazaki, Shun; Hirano, Ikuo; Sekine, Hiroki; Dan, Takashi; Takeda, Kotaro; Fong, Guo-Hua; Nishiyama, Akira; Ito, Sadayoshi; Miyata, Toshio; Yamamoto, Masayuki; Suzuki, Norio

    2016-02-01

    Erythropoietin (Epo) is produced by renal Epo-producing cells (REPs) in a hypoxia-inducible manner. The conversion of REPs into myofibroblasts and coincident loss of Epo-producing ability are the major cause of renal fibrosis and anemia. However, the hypoxic response of these transformed myofibroblasts remains unclear. Here, we used complementary in vivo transgenic and live imaging approaches to better understand the importance of hypoxia signaling in Epo production. Live imaging of REPs in transgenic mice expressing green fluorescent protein from a modified Epo-gene locus revealed that healthy REPs tightly associated with endothelium by wrapping processes around capillaries. However, this association was hampered in states of renal injury-induced inflammation previously shown to correlate with the transition to myofibroblast-transformed renal Epo-producing cells (MF-REPs). Furthermore, activation of hypoxia-inducible factors (HIFs) by genetic inactivation of HIF-prolyl hydroxylases (PHD1, PHD2, and PHD3) selectively in Epo-producing cells reactivated Epo production in MF-REPs. Loss of PHD2 in REPs restored Epo-gene expression in injured kidneys but caused polycythemia. Notably, combined deletions of PHD1 and PHD3 prevented loss of Epo expression without provoking polycythemia. Mice with PHD-deficient REPs also showed resistance to LPS-induced Epo repression in kidneys, suggesting that augmented HIF signaling counterbalances inflammatory stimuli in regulation of Epo production. Thus, augmentation of HIF signaling may be an attractive therapeutic strategy for treating renal anemia by reactivating Epo synthesis in MF-REPs. PMID:26054543

  8. Plasma soluble erythropoietin receptor is decreased during sleep in Andean highlanders with Chronic Mountain Sickness

    PubMed Central

    Corante, Noemí; Anza-Ramírez, Cecilia; Figueroa-Mujíca, Rómulo; Vizcardo-Galindo, Gustavo; Mercado, Andy; Macarlupú, José Luis; León-Velarde, Fabiola

    2016-01-01

    Excessive erythrocytosis (EE) is the main sign of Chronic Mountain Sickness (CMS), a highly prevalent syndrome in Andean highlanders. Low pulse O2 saturation (SpO2) during sleep and serum androgens have been suggested to contribute to EE in CMS patients. However, whether these factors have a significant impact on the erythropoietin (Epo) system leading to EE is still unclear. We have recently shown that morning soluble Epo receptor (sEpoR), an endogenous Epo antagonist, is decreased in CMS patients suggesting increased Epo availability (increased Epo/sEpoR). The present study aimed to characterize the nocturnal concentration profile of sEpoR and Epo and their relationship with SpO2, Hct, and serum testosterone in healthy highlanders (HH) and CMS patients. Epo and sEpoR concentrations were evaluated every 4 h (6 PM to 6 AM) and nighttime SpO2 was continuously monitored (10 PM to 6 AM) in 39 male participants (CMS, n = 23; HH, n = 16) aged 21-65 yr from Cerro de Pasco, Peru (4,340 m). CMS patients showed higher serum Epo concentrations throughout the night and lower sEpoR from 10 PM to 6 AM. Consequently, Epo/sEpoR was significantly higher in the CMS group at every time point. Mean sleep-time SpO2 was lower in CMS patients compared with HH, while the percentage of sleep time spent with SpO2 < 80% was higher. Multiple-regression analysis showed mean sleep-time SpO2 and Epo/sEpoR as significant predictors of hematocrit corrected for potential confounders (age, body mass index, and testosterone). Testosterone levels were associated neither with Hct nor with erythropoietic factors. In conclusion, our results show sustained erythropoietic stimulus driven by the Epo system in CMS patients, further enhanced by a continuous exposure to accentuated nocturnal hypoxemia. PMID:27125843

  9. Plasma soluble erythropoietin receptor is decreased during sleep in Andean highlanders with Chronic Mountain Sickness.

    PubMed

    Villafuerte, Francisco C; Corante, Noemí; Anza-Ramírez, Cecilia; Figueroa-Mujíca, Rómulo; Vizcardo-Galindo, Gustavo; Mercado, Andy; Macarlupú, José Luis; León-Velarde, Fabiola

    2016-07-01

    Excessive erythrocytosis (EE) is the main sign of Chronic Mountain Sickness (CMS), a highly prevalent syndrome in Andean highlanders. Low pulse O2 saturation (SpO2) during sleep and serum androgens have been suggested to contribute to EE in CMS patients. However, whether these factors have a significant impact on the erythropoietin (Epo) system leading to EE is still unclear. We have recently shown that morning soluble Epo receptor (sEpoR), an endogenous Epo antagonist, is decreased in CMS patients suggesting increased Epo availability (increased Epo/sEpoR). The present study aimed to characterize the nocturnal concentration profile of sEpoR and Epo and their relationship with SpO2, Hct, and serum testosterone in healthy highlanders (HH) and CMS patients. Epo and sEpoR concentrations were evaluated every 4 h (6 PM to 6 AM) and nighttime SpO2 was continuously monitored (10 PM to 6 AM) in 39 male participants (CMS, n = 23; HH, n = 16) aged 21-65 yr from Cerro de Pasco, Peru (4,340 m). CMS patients showed higher serum Epo concentrations throughout the night and lower sEpoR from 10 PM to 6 AM. Consequently, Epo/sEpoR was significantly higher in the CMS group at every time point. Mean sleep-time SpO2 was lower in CMS patients compared with HH, while the percentage of sleep time spent with SpO2 < 80% was higher. Multiple-regression analysis showed mean sleep-time SpO2 and Epo/sEpoR as significant predictors of hematocrit corrected for potential confounders (age, body mass index, and testosterone). Testosterone levels were associated neither with Hct nor with erythropoietic factors. In conclusion, our results show sustained erythropoietic stimulus driven by the Epo system in CMS patients, further enhanced by a continuous exposure to accentuated nocturnal hypoxemia.

  10. Erythropoietin Does Not Enhance Skeletal Muscle Protein Synthesis Following Exercise in Young and Older Adults

    PubMed Central

    Lamon, Séverine; Zacharewicz, Evelyn; Arentson-Lantz, Emily; Gatta, Paul A. Della; Ghobrial, Lobna; Gerlinger-Romero, Frederico; Garnham, Andrew; Paddon-Jones, Douglas; Russell, Aaron P.

    2016-01-01

    Purpose: Erythropoietin (EPO) is a renal cytokine that is primarily involved in hematopoiesis while also playing a role in non-hematopoietic tissues expressing the EPO-receptor (EPOR). The EPOR is present in human skeletal muscle. In mouse skeletal muscle, EPO stimulation can activate the AKT serine/threonine kinase 1 (AKT) signaling pathway, the main positive regulator of muscle protein synthesis. We hypothesized that a single intravenous EPO injection combined with acute resistance exercise would have a synergistic effect on skeletal muscle protein synthesis via activation of the AKT pathway. Methods: Ten young (24.2 ± 0.9 years) and 10 older (66.6 ± 1.1 years) healthy subjects received a primed, constant infusion of [ring-13C6] L-phenylalanine and a single injection of 10,000 IU epoetin-beta or placebo in a double-blind randomized, cross-over design. 2 h after the injection, the subjects completed an acute bout of leg extension resistance exercise to stimulate skeletal muscle protein synthesis. Results: Significant interaction effects in the phosphorylation levels of the members of the AKT signaling pathway indicated a differential activation of protein synthesis signaling in older subjects when compared to young subjects. However, EPO offered no synergistic effect on vastus lateralis mixed muscle protein synthesis rate in young or older subjects. Conclusions: Despite its ability to activate the AKT pathway in skeletal muscle, an acute EPO injection had no additive or synergistic effect on the exercise-induced activation of muscle protein synthesis or muscle protein synthesis signaling pathways. PMID:27458387

  11. Erythropoietin enhancement of rat pancreatic tumor cell proliferation requires the activation of ERK and JNK signals.

    PubMed

    Bose, Chhanda; Udupa, Kodetthoor B

    2008-08-01

    Erythropoietin (EPO) regulates the proliferation and differentiation of erythroid cells by binding to its specific transmembrane receptor EPOR. Recent studies, however, have shown that the EPOR is additionally present in various cancer cells and EPO induces the proliferation of these cells, suggesting a different function for EPO other than erythropoiesis. Therefore, the purpose of the present study was to examine EPOR expression and the role of EPO in the proliferation and signaling cascades involved in this process, using the rat pancreatic tumor cell line AR42J. Our results showed that AR42J cells expressed EPOR, and EPO significantly enhanced their proliferation. Cell cycle analysis of EPO-treated cells indicated an increased percentage of cells in the S phase, whereas cell numbers in G0/G1 phase were significantly reduced. Phosphorylation of extracellular regulatory kinase 1/2 (ERK1/2) and c-Jun NH(2) terminal kinase 1/2 (JNK1/2) was rapidly stimulated and sustained after EPO addition. Treatment of cells with mitogen-activated protein/ERK kinase (MEK) inhibitor PD98059 or JNK inhibitor SP600125 significantly inhibited EPO-enhanced proliferation and also increased the fraction of cells in G0/G1 phase. Furthermore, the inhibition of JNK using small interference RNA (siRNA) suppressed EPO-enhanced proliferation of AR42J cells. Taken together, our results indicate that AR42J cells express EPOR and that the activation of both ERK1/2 and JNK1/2 by EPO is essential in regulating proliferation and the cell cycle. Thus both appear to play a key role in EPO-enhanced proliferation and suggest that the presence of both is required for EPO-mediated proliferation of AR42J cells.

  12. Effects of Recombinant Human Erythropoietin on Resistance Artery Endothelial Function in Stage 4 Chronic Kidney Disease

    PubMed Central

    Briet, Marie; Barhoumi, Tlili; Mian, Muhammad Oneeb Rehman; Sierra, Cristina; Boutouyrie, Pierre; Davidman, Michael; Bercovitch, David; Nessim, Sharon J.; Frisch, Gershon; Paradis, Pierre; Lipman, Mark L.; Schiffrin, Ernesto L.

    2013-01-01

    Background Recent studies have raised concern about the safety of erythropoiesis‐stimulating agents because of evidence of increased risk of hypertension and cardiovascular morbidity and mortality in chronic kidney disease (CKD) patients. In the present study, we investigated the effects of recombinant human erythropoietin (EPO) on endothelial function of gluteal subcutaneous resistance arteries isolated from 17 stage 4 patients (estimated glomerular filtration rate 21.9±7.4 mL/min per 1.73 m2) aged 63±13 years. Methods and Results Arteries were mounted on a pressurized myograph. EPO impaired endothelium‐dependent relaxation in a concentration‐dependent manner. The maximal response to acetylcholine with EPO at 1, 10, and 20 IU/mL was reduced by 12%, 34%, and 43%, respectively, compared with the absence of EPO (P<0.001). EPO‐induced endothelial dysfunction was significantly associated with carotid stiffness and history of cardiovascular events. EPO had no effect on norepinephrine‐induced vasoconstriction or sodium nitroprusside–induced relaxation. ABT‐627, an endothelin type A receptor antagonist, and tempol, a superoxide dismutase mimetic, partially reversed the altered endothelial function in the presence of EPO (P<0.01). Increased expression of endothelin‐1 was found in the vessel wall after incubation with EPO. Conclusions EPO alters endothelial function of resistance arteries in CKD patients via a mechanism involving in part oxidative stress and signaling through an endothelin type A receptor. EPO‐induced endothelial dysfunction could contribute to deleterious effects of EPO described in large interventional trials. PMID:23584809

  13. Doping with growth hormone/IGF-1, anabolic steroids or erythropoietin: is there a cancer risk?

    PubMed

    Tentori, Lucio; Graziani, Grazia

    2007-05-01

    Anabolic steroid and peptide hormones or growth factors are utilized to increase the performance of athletes of professional or amateur sports. Despite their well-documented adverse effects, the use of some of these agents has significantly grown and has been extended also to non-athletes with the aim to improve appearance or to counteract ageing. Pre-clinical studies and epidemiological observations in patients with an excess of hormone production or in patients chronically treated with hormones/growth factors for various pathologies have warned about the potential risk of cancer development and progression which may be also associated to the use of certain doping agents. Anabolic steroids have been described to provoke liver tumours; growth hormone or high levels of its mediator insulin-like growth factor-1 (IGF-1) have been associated with colon, breast, and prostate cancers. Actually, IGF-1 promotes cell cycle progression and inhibits apoptosis either by triggering other growth factors or by interacting with pathways which have an established role in carcinogenesis and cancer promotion. More recently, the finding that erythropoietin (Epo) may promote angiogenesis and inhibit apoptosis or modulate chemo- or radiosensitivity in cancer cells expressing the Epo receptor, raised the concern that the use of recombinant Epo to increase tissue oxygenation might favour tumour survival and aggressiveness. Cancer risk associated to doping might be higher than that of patients using hormones/growth factors as replacement therapy, since enormous doses are taken by the athletes often for a long period of time. Moreover, these substances are often used in combination with other licit or illicit drugs and this renders almost unpredictable all the possible adverse effects including cancer. Anyway, athletes should be made aware that long-term treatment with doping agents might increase the risk of developing cancer.

  14. Role of ATP-dependent K channels in the effects of erythropoietin in renal ischaemia injury

    PubMed Central

    Yilmaz, Tonguç Utku; Yazihan, Nuray; Dalgic, Aydın; Kaya, Ezgi Ermis; Salman, Bulent; Kocak, Mehtap; Akcil, Ethem

    2015-01-01

    Background & objectives: Erythropoietin (EPO) has cytoprotective and anti-apoptotic effects in pathological conditions, including hypoxia and ischaemia-reperfusion injury. One of the targets to protect against injury is ATP-dependent potassium (KATP) channels. These channels could be involved in EPO induced ischaemic preconditoning like a protective effect. We evaluated the cell cytoprotective effects of EPO in relation to KATP channel activation in the renal tubular cell culture model under hypoxic/normoxic conditions. Methods: Dose and time dependent effects of EPO, KATP channel blocker glibenclamide and KATP channel opener diazoxide on cellular proliferation were evaluated by colorimetric assay MTT [3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide] under normoxic and hypoxic conditions in human renal proximal tubular cell line (CRL-2830). Evaluation of the dose and time dependent effects of EPO, glibenclamide and diazoxide on apoptosis was done by caspase-3 activity levels. Hypoxia inducible factor-1 alpha (HIF-1 α) mRNA levels were measured by semi-quantative reverse transcription polymerase chain reaction (RT)-PCR. Kir 6.1 protein expresion was evalutaed by Western blot. Results: Glibenclamide treatment decreased the number of living cells in a time and dose dependent manner, whereas EPO and diazoxide treatments increased. Glibenclamide (100 μM) treatment significantly blocked the anti-apoptotic effects of EPO (10 IU/ml) under both normoxic and hypoxic conditions. EPO (10 IU/ml) and diazoxide (100 μM) treatments significantly increased (P<0.01) whereas glibenclamide decreased (P<0.05) HIF-1 α mRNA expression. Glibenclamide significantly (P<0.01) decreased EPO induced HIF-1 α mRNA expression when compared with the EPO alone group. Interpretation & conclusions: Our results showed that the cell proliferative, cytoprotective and anti-apoptotic effects of EPO were associated with KATP channels in the renal tubular cell culture model under hypoxic

  15. EPOR-Based Purification and Analysis of Erythropoietin Mimetic Peptides from Human Urine by Cys-Specific Cleavage and LC/MS/MS

    NASA Astrophysics Data System (ADS)

    Vogel, Matthias; Thomas, Andreas; Schänzer, Wilhelm; Thevis, Mario

    2015-09-01

    The development of a new class of erythropoietin mimetic agents (EMA) for treating anemic conditions has been initiated with the discovery of oligopeptides capable of dimerizing the erythropoietin (EPO) receptor and thus stimulating erythropoiesis. The most promising amino acid sequences have been mounted on various different polymeric structures or carrier molecules to obtain highly active EPO-like drugs exhibiting beneficial and desirable pharmacokinetic profiles. Concomitant with creating new therapeutic options, erythropoietin mimetic peptide (EMP)-based drug candidates represent means to artificially enhance endurance performance and necessitate coverage by sports drug testing methods. Therefore, the aim of the present study was to develop a strategy for the comprehensive detection of EMPs in doping controls, which can be used complementary to existing protocols. Three model EMPs were used to provide proof-of-concept data. Following EPO receptor-facilitated purification of target analytes from human urine, the common presence of the cysteine-flanked core structure of EMPs was exploited to generate diagnostic peptides with the aid of a nonenzymatic cleavage procedure. Sensitive detection was accomplished by targeted-SIM/data-dependent MS2 analysis. Method characterization was conducted for the EMP-based drug peginesatide concerning specificity, linearity, precision, recovery, stability, ion suppression/enhancement, and limit of detection (LOD, 0.25 ng/mL). Additionally, first data for the identification of the erythropoietin mimetic peptides EMP1 and BB68 were generated, demonstrating the multi-analyte testing capability of the presented approach.

  16. Evolving insights into the synergy between erythropoietin and thrombopoietin and the bipotent erythroid/megakaryocytic progenitor cell.

    PubMed

    Papayannopoulou, Thalia; Kaushansky, Kenneth

    2016-08-01

    Although the synergy between erythropoietin and thrombopoietin has previously been pointed out, the clonal demonstration of a human bipotent erythroid/megakaryocytic progenitor (MEP) was first published in Experimental Hematology (Papayannopoulou T, Brice M, Farrer D, Kaushansky K. Exp Hematol. 1996;24:660-669) and later in the same year in Blood (Debili N, Coulombel L, Croisille L, et al. Blood. 1996;88:1284-1296). This demonstration, and the fact that both bipotent and monopotent erythroid or megakaryocytic progenitors co-express markers of both lineages and respond to both lineage-specific transcription factors, has provided a background for the extensive use of MEP assessment by fluorescence-activated cell sorting in many subsequent studies. Beyond this, the demonstration of shared regulatory elements and the presence of single mutations affecting both lineages have inspired further studies to decipher how the shift in transcription factor networks occurs from one lineage to the other. Furthermore, in addition to shared effects, erythropoietin and thrombopoietin have additional independent effects. Most notable for thrombopoietin is its effect on hematopoietic stem cells illustrated by in vitro and in vivo approaches. PMID:26773569

  17. ARA 290, a Nonerythropoietic Peptide Engineered from Erythropoietin, Improves Metabolic Control and Neuropathic Symptoms in Patients with Type 2 Diabetes

    PubMed Central

    Brines, Michael; Dunne, Ann N; van Velzen, Monique; Proto, Paolo L; Ostenson, Claes-Goran; Kirk, Rita I; Petropoulos, Ioannis N; Javed, Saad; Malik, Rayaz A; Cerami, Anthony; Dahan, Albert

    2014-01-01

    Although erythropoietin ameliorates experimental type 2 diabetes with neuropathy, serious side effects limit its potential clinical use. ARA 290, a nonhematopoietic peptide designed from the structure of erythropoietin, interacts selectively with the innate repair receptor that mediates tissue protection. ARA 290 has shown efficacy in preclinical and clinical studies of metabolic control and neuropathy. To evaluate the potential activity of ARA 290 in type 2 diabetes and painful neuropathy, subjects were enrolled in this phase 2 study. ARA 290 (4 mg) or placebo were self-administered subcutaneously daily for 28 d and the subjects followed for an additional month without further treatment. No potential safety issues were identified. Subjects receiving ARA 290 exhibited an improvement in hemoglobin A1c (Hb A1c) and lipid profiles throughout the 56 d observation period. Neuropathic symptoms as assessed by the PainDetect questionnaire improved significantly in the ARA 290 group. Mean corneal nerve fiber density (CNFD) was reduced significantly compared with normal controls and subjects with a mean CNFD >1 standard deviation from normal showed a significant increase in CNFD compared with no change in the placebo group. These observations suggest that ARA 290 may benefit both metabolic control and neuropathy in subjects with type 2 diabetes and deserves continued clinical evaluation. PMID:25387363

  18. Establishment and characterization of a new erythropoietin-dependent acute myeloid leukemia cell line, AS-E2.

    PubMed

    Miyazaki, Y; Kuriyama, K; Higuchi, M; Tsushima, H; Sohda, H; Imai, N; Saito, M; Kondo, T; Tomonaga, M

    1997-11-01

    We have established an erythropoietin-dependent human leukemia cell line, AS-E2, from a patient with acute myeloid leukemia. These cells have many characteristics of late erythroid progenitor cells, they are positive for CD36, Glycophorin A, and CD71 but negative for CD41, and positive for benzidine and PAS staining. These cells express GATA-1 and have low affinity erythropoietin (EPO) receptor on their surface. Interestingly, AS-E2 cells are strictly dependent on EPO for their growth and survival; other cytokines including GM-CSF, stem cell factor, or IL-3 cannot support the growth of this cell line. These features are similar to late erythroid lineage cells, like normal BFU-E or CFU-E, and we have demonstrated that EPO stimulation induces the tyrosine phosphorylation of several proteins in AS-E2 cells including the EPO receptor and JAK2 kinase. This new cell line is a useful reagent to study biological and molecular events during the late stages of erythropoiesis, and to understand transforming events in human erythroid cells.

  19. Far-western blotting as a solution to the non-specificity of the anti-erythropoietin receptor antibody

    PubMed Central

    Fecková, Barbora; Kimáková, Patrícia; Ilkovičová, Lenka; Szentpéteriová, Erika; Debeljak, Nataša; Solárová, Zuzana; Sačková, Veronika; Šemeláková, Martina; Bhide, Mangesh; Solár, Peter

    2016-01-01

    The erythropoietin receptor (EpoR) is a member of the cytokine receptor family. The interaction between erythropoietin (Epo) and EpoR is important for the production and maturation of erythroid cells, resulting in the stimulation of hematopoiesis. The fact that EpoR was also detected in neoplastic cells has opened the question about the relevance of anemia treatment with recombinant Epo in cancer patients. Numerous studies have reported pro-stimulating and anti-apoptotic effects of Epo in cancer cells, thus demonstrating EpoR functionality in these cells. By contrast, a previous study claims the absence of EpoR in tumor cells. This apparent discrepancy is based, according to certain authors, on the use of non-specific anti-EpoR antibodies. With the aim of bypassing the direct detection of EpoR with an anti-EpoR antibody, the present authors propose a far-western blot methodology, which in addition, confirms the interaction of Epo with EpoR. Applying this technique, the presence of EpoR and its interaction with Epo in human ovarian adenocarcinoma A2780 and normal human umbilical vein endothelial cells was confirmed. Furthermore, modified immunoprecipitation of EpoR followed by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry analysis confirmed a 57 kDa protein as a human Epo-interacting protein in both cell lines. PMID:27446474

  20. Methylation impact analysis of erythropoietin (EPO) Gene to hypoxia inducible factor-1α (HIF-1α) activity.

    PubMed

    Dewi, Firli Rahmah Primula; Fatchiyah, Fatchiyah

    2013-01-01

    Erythropoietin (EPO) is a glycoprotein hormone that play a role as key regulator in the production of red blood cells. The promoter region of EPO is methylated in normoxic (non-hypoxia) condition, but not in hypoxic condition. Methylation of the EPO enhancer region decline the transcription activity of EPO gene. The aim of this study is to investigate how different methylation percentage affected on the regulation and transcriptional activity of EPO gene. The DNA sequence of erythropoietin gene and protein sequence was retrieved from the sequence database of NCBI. DNA structure was constructed using 3D-DART web server and modeling structure of HIF1 predicted using SWISS-MODEL web server. Methylated DNA sequence of EPO gene using performed with YASARA View software and docking of EPO gene and transcription factor HIF1 analyzed by using HADDOCK webserver. Our result showed that binding energy in 46% methylated DNA was higher (-161,45 kcal/mol) than in unmethylated DNA (-194,16 kcal/mol) and 8% methylated DNA (-175,94 kcal/mol). So, we presume that a silencing mechanism of the Epo gene by methylation is correlated with the binding energy, which is required for interaction. A higher methylation percentage correlates with a higher binding energy which can cause an unstable interaction between DNA and transcription factor. In conclution, methylation of promoter and enhancer region of Epo gene leads to silencing. PMID:24023421

  1. Senior Administrators Should Have Administrative Contracts.

    ERIC Educational Resources Information Center

    Posner, Gary J.

    1987-01-01

    Recognizing that termination is viewed by the employee as the equivalent to capital punishment of a career, an administrative contract can reduce the emotional and financial entanglements that often result. Administrative contracts are described. (MLW)

  2. Elevated Endogenous Erythropoietin Concentrations Are Associated with Increased Risk of Brain Damage in Extremely Preterm Neonates

    PubMed Central

    Korzeniewski, Steven J.; Allred, Elizabeth; Logan, J. Wells; Fichorova, Raina N.; Engelke, Stephen; Kuban, Karl C. K.; O’Shea, T. Michael; Paneth, Nigel; Holm, Mari; Dammann, Olaf; Leviton, Alan

    2015-01-01

    Background We sought to determine, in very preterm infants, whether elevated perinatal erythropoietin (EPO) concentrations are associated with increased risks of indicators of brain damage, and whether this risk differs by the co-occurrence or absence of intermittent or sustained systemic inflammation (ISSI). Methods Protein concentrations were measured in blood collected from 786 infants born before the 28th week of gestation. EPO was measured on postnatal day 14, and 25 inflammation-related proteins were measured weekly during the first 2 postnatal weeks. We defined ISSI as a concentration in the top quartile of each of 25 inflammation-related proteins on two separate days a week apart. Hypererythropoietinemia (hyperEPO) was defined as the highest quartile for gestational age on postnatal day 14. Using logistic regression and multinomial logistic regression models, we compared risks of brain damage among neonates with hyperEPO only, ISSI only, and hyperEPO+ISSI, to those who had neither hyperEPO nor ISSI, adjusting for gestational age. Results Newborns with hyperEPO, regardless of ISSI, were more than twice as likely as those without to have very low (< 55) Mental (OR 2.3; 95% CI 1.5-3.5) and/or Psychomotor (OR 2.4; 95% CI 1.6-3.7) Development Indices (MDI, PDI), and microcephaly at age two years (OR 2.4; 95%CI 1.5-3.8). Newborns with both hyperEPO and ISSI had significantly increased risks of ventriculomegaly, hemiparetic cerebral palsy, microcephaly, and MDI and PDI < 55 (ORs ranged from 2.2-6.3), but not hypoechoic lesions or other forms of cerebral palsy, relative to newborns with neither hyperEPO nor ISSI. Conclusion hyperEPO, regardless of ISSI, is associated with elevated risks of very low MDI and PDI, and microcephaly, but not with any form of cerebral palsy. Children with both hyperEPO and ISSI are at higher risk than others of very low MDI and PDI, ventriculomegaly, hemiparetic cerebral palsy, and microcephaly. PMID:25793991

  3. Effects of ammonia on CHO cell growth, erythropoietin production, and glycosylation.

    PubMed

    Yang, M; Butler, M

    2000-05-20

    The effect of ammonium chloride was determined on a culture of CHO cells transfected with the human erythropoietin (EPO) gene. Cell growth was inhibited above a culture concentration of 5 mM NH(4)Cl with an IC-50 determined to be 33 mM. The specific production of EPO increased with the addition of NH(4)Cl above 5 mM. At 10 mM NH(4)Cl, the final cell density after 4 days in culture was significantly lower but the final yield of EPO was significantly higher. This appeared to be due to continued protein production after cell growth had ceased. The metabolic effects of added NH(4)Cl included higher specific consumption rates of glucose and glutamine and an increased rate of production of alanine, glycine, and glutamate. The EPO analyzed from control cultures had a molecular weight range of 33-39 kDa and an isoelectric point range of 4.06-4.67. Seven distinct isoforms of the molecule were identified by two-dimensional electrophoresis. This molecular heterogeneity was ascribed to variable glycosylation. Complete enzymatic de-glycosylation resulted in a single molecular form with a molecular mass of 18 kDa. Addition of NH(4)Cl to the cultures caused a significant increase in the heterogeneity of the glycoforms as shown by an increased molecular weight and pI range. Enzymatic de-sialylation of the EPO from the ammonia-treated and control cultures resulted in identical electrophoretic patterns. This indicated that the effect of ammonia was in the reduction of terminal sialylation of the glycan structures which accounted for the increased pI. Selective removal of the N-glycan structures by PNGase F resulted in two bands identified as the O-glycan linked structure (19 kDa) and the completely de-glycosylated structure (18 kDa). The proportion of the O-linked glycan structure was reduced, and its pI increased in cultures to which ammonia was added. Thus, the glycosylation pattern altered by the presence of ammonia included a reduction in terminal sialylation of all the glycans

  4. Erythropoietin in amyotrophic lateral sclerosis: a multicentre, randomised, double blind, placebo controlled, phase III study

    PubMed Central

    Lauria, Giuseppe; Dalla Bella, Eleonora; Antonini, Giovanni; Borghero, Giuseppe; Capasso, Margherita; Caponnetto, Claudia; Chiò, Adriano; Corbo, Massimo; Eleopra, Roberto; Fazio, Raffaella; Filosto, Massimiliano; Giannini, Fabio; Granieri, Enrico; La Bella, Vincenzo; Logroscino, Giancarlo; Mandrioli, Jessica; Mazzini, Letizia; Monsurrò, Maria Rosaria; Mora, Gabriele; Pietrini, Vladimiro; Quatrale, Rocco; Rizzi, Romana; Salvi, Fabrizio; Siciliano, Gabriele; Sorarù, Gianni; Volanti, Paolo; Tramacere, Irene; Filippini, Graziella

    2015-01-01

    Objective To assess the efficacy of recombinant human erythropoietin (rhEPO) in amyotrophic lateral sclerosis (ALS). Methods Patients with probable laboratory-supported, probable or definite ALS were enrolled by 25 Italian centres and randomly assigned (1:1) to receive intravenous rhEPO 40 000 IU or placebo fortnightly as add-on treatment to riluzole 100 mg daily for 12 months. The primary composite outcome was survival, tracheotomy or >23 h non-invasive ventilation (NIV). Secondary outcomes were ALSFRS-R, slow vital capacity (sVC) and quality of life (ALSAQ-40) decline. Tolerability was evaluated analysing adverse events (AEs) causing withdrawal. The randomisation sequence was computer-generated by blocks, stratified by centre, disease severity (ALSFRS-R cut-off score of 33) and onset (spinal or bulbar). The main outcome analysis was performed in all randomised patients and by intention-to-treat for the entire population and patients stratified by severity and onset. The study is registered, EudraCT 2009-016066-91. Results We randomly assigned 208 patients, of whom 5 (1 rhEPO and 4 placebo) withdrew consent and 3 (placebo) became ineligible (retinal thrombosis, respiratory insufficiency, SOD1 mutation) before receiving treatment; 103 receiving rhEPO and 97 placebo were eligible for analysis. At 12 months, the annualised rate of death (rhEPO 0.11, 95% CI 0.06 to 0.20; placebo: 0.08, CI 0.04 to 0.17), tracheotomy or >23 h NIV (rhEPO 0.16, CI 0.10 to 0.27; placebo 0.18, CI 0.11 to 0.30) did not differ between groups, also after stratification by onset and ALSFRS-R at baseline. Withdrawal due to AE was 16.5% in rhEPO and 8.3% in placebo. No differences were found for secondary outcomes. Conclusions RhEPO 40 000 IU fortnightly did not change the course of ALS. PMID:25595151

  5. Erythropoietin improves skeletal muscle microcirculation and tissue bioenergetics in a mouse sepsis model

    PubMed Central

    Kao, Raymond; Xenocostas, Anargyros; Rui, Tao; Yu, Pei; Huang, Weixiong; Rose, James; Martin, Claudio M

    2007-01-01

    Introduction The relationship between oxygen delivery and consumption in sepsis is impaired, suggesting a microcirculatory perfusion defect. Recombinant human erythropoietin (rHuEPO) regulates erythropoiesis and also exerts complex actions promoting the maintenance of homeostasis of the organism under stress. The objective of this study was to test the hypothesis that rHuEPO could improve skeletal muscle capillary perfusion and tissue oxygenation in sepsis. Methods Septic mice in three experiments received rHu-EPO 400 U/kg subcutaneously 18 hours after cecal ligation and perforation (CLP). The first experiment measured the acute effects of rHuEPO on hemodynamics, blood counts, and arterial lactate level. The next two sets of experiments used intravital microscopy to observe capillary perfusion and nicotinamide adenine dinucleotide (NADH) fluorescence post-CLP after treatment with rHuEPO every 10 minutes for 40 minutes and at 6 hours. Perfused capillary density during a three-minute observation period and NADH fluorescence were measured. Results rHuEPO did not have any effects on blood pressure, lactate level, or blood cell numbers. CLP mice demonstrated a 22% decrease in perfused capillary density compared to the sham group (28.5 versus 36.6 capillaries per millimeter; p < 0.001). Treatment of CLP mice with rHuEPO resulted in an immediate and significant increase in perfused capillaries in the CLP group at all time points compared to baseline from 28.5 to 33.6 capillaries per millimeter at 40 minutes; p < 0.001. A significant increase in baseline NADH, suggesting tissue hypoxia, was noted in the CLP mice compared to the sham group (48.3 versus 43.9 fluorescence units [FU]; p = 0.03) and improved with rHuEPO from 48.3 to 44.4 FU at 40 minutes (p = 0.02). Six hours after treatment with rHuEPO, CLP mice demonstrated a higher mean perfused capillary density (39.4 versus 31.7 capillaries per millimeter; p < 0.001) and a lower mean NADH fluorescence as compared to CLP

  6. Determinants of erythropoietin release in response to short-term hypobaric hypoxia

    NASA Technical Reports Server (NTRS)

    Ge, Ri-Li; Witkowski, S.; Zhang, Y.; Alfrey, C.; Sivieri, M.; Karlsen, T.; Resaland, G. K.; Harber, M.; Stray-Gundersen, J.; Levine, B. D.

    2002-01-01

    We measured blood erythropoietin (EPO) concentration, arterial O(2) saturation (Sa(O(2))), and urine PO(2) in 48 subjects (32 men and 16 women) at sea level and after 6 and 24 h at simulated altitudes of 1,780, 2,085, 2,454, and 2,800 m. Renal blood flow (Doppler) and Hb were determined at sea level and after 6 h at each altitude (n = 24) to calculate renal O(2) delivery. EPO increased significantly after 6 h at all altitudes and continued to increase after 24 h at 2,454 and 2,800 m, although not at 1,780 or 2,085 m. The increase in EPO varied markedly among individuals, ranging from -41 to 400% after 24 h at 2,800 m. Similar to EPO, urine PO(2) decreased after 6 h at all altitudes and returned to baseline by 24 h at the two lowest altitudes but remained decreased at the two highest altitudes. Urine PO(2) was closely related to EPO via a curvilinear relationship (r(2) = 0.99), although also with prominent individual variability. Renal blood flow remained unchanged at all altitudes. Sa(O(2)) decreased slightly after 6 h at the lowest altitudes but decreased more prominently at the highest altitudes. There were only modest, albeit statistically significant, relationships between EPO and Sa(O(2)) (r = 0.41, P < 0.05) and no significant relationship with renal O(2) delivery. These data suggest that 1) the altitude-induced increase in EPO is "dose" dependent: altitudes > or =2,100-2,500 m appear to be a threshold for stimulating sustained EPO release in most subjects; 2) short-term acclimatization may restore renal tissue oxygenation and restrain the rise in EPO at the lowest altitudes; and 3) there is marked individual variability in the erythropoietic response to altitude that is only partially explained by "upstream" physiological factors such as those reflecting O(2) delivery to EPO-producing tissues.

  7. Sequential univariate gating approach to study the effects of erythropoietin in murine bone marrow.

    PubMed

    Achuthanandam, Ram; Quinn, John; Capocasale, Renold J; Bugelski, Peter J; Hrebien, Leonid; Kam, Moshe

    2008-08-01

    Analysis of multicolor flow cytometric data is traditionally based on the judgment of an expert, generally time consuming, sometimes incomplete and often subjective in nature. In this article, we investigate another statistical method using a Sequential Univariate Gating (SUG) algorithm to identify regions of interest between two groups of multivariate flow cytometric data. The metric used to differentiate between the groups of univariate distributions in SUG is the Kolmogorov-Smirnov distance (D) statistic. The performance of the algorithm is evaluated by applying it to a known three-color data set looking at activation of CD4+ and CD8+ lymphocytes with anti-CD3 antibody treatment and comparing the results to the expert analysis. The algorithm is then applied to a four-color data set used to study the effects of recombinant human erythropoietin (rHuEPO) on several murine bone marrow populations. SUG was used to identify regions of interest in the data and results compared to expert analysis and the current state-of-the-art statistical method, Frequency Difference Gating (FDG). Cluster analysis was then performed to identify subpopulations responding differently to rHuEPO. Expert analysis, SUG and FDG identified regions in the data that showed activation of CD4+ and CD8+ lymphocytes with anti-CD3 treatment. In the rHuEPO treated data sets, the expert and SUG identified a dose responsive expansion of only the erythroid precursor population. In contrast, FDG resulted in identification of regions of interest both in the erythroid precursors as well as in other bone marrow populations. Clustering within the regions of interest defined by SUG resulted in identification of four subpopulations of erythroid precursors that are morphologically distinct and show a differential response to rHuEPO treatment. Greatest expansion is seen in the basophilic and poly/orthochromic erythroblast populations with treatment. Identification of populations of interest can be performed

  8. Erythropoietin does not reduce plasma lactate, H⁺, and K⁺ during intense exercise.

    PubMed

    Nordsborg, N B; Robach, P; Boushel, R; Calbet, J A L; Lundby, C

    2015-12-01

    It is investigated if recombinant human erythropoietin (rHuEPO) treatment for 15 weeks (n = 8) reduces extracellular accumulation of metabolic stress markers such as lactate, H(+) , and K(+) during incremental exhaustive exercise. After rHuEPO treatment, normalization of blood volume and composition by hemodilution preceded an additional incremental test. Group averages were calculated for an exercise intensity ∼80% of pre-rHuEPO peak power output. After rHuEPO treatment, leg lactate release to the plasma compartment was similar to before (4.3 ± 1.6 vs 3.9 ± 2.5 mmol/min) and remained similar after hemodilution. Venous lactate concentration was higher (P < 0.05) after rHuEPO treatment (7.1 ± 1.6 vs 5.2 ± 2.1 mM). Leg H(+) release to the plasma compartment after rHuEPO was similar to before (19.6 ± 5.4 vs 17.6 ± 6.0 mmol/min) and remained similar after hemodilution. Nevertheless, venous pH was lower (P < 0.05) after rHuEPO treatment (7.18 ± 0.04 vs 7.22 ± 0.05). Leg K(+) release to the plasma compartment after rHuEPO treatment was similar to before (0.8 ± 0.5 vs 0.7 ± 0.7 mmol/min) and remained similar after hemodilution. Additionally, venous K(+) concentrations were similar after vs before rHuEPO (5.3 ± 0.3 vs 5.1 ± 0.4 mM). In conclusion, rHuEPO does not reduce plasma accumulation of lactate, H(+) , and K(+) at work rates corresponding to ∼80% of peak power output. PMID:25556620

  9. Enhancement of recombinant erythropoietin production in CHO cells in an incubator without CO(2) addition.

    PubMed

    Yoon, S K; Ahn, Y H; Han, K

    2001-10-01

    The effect of low levels of carbon dioxide (CO(2)) in the gas phase on the production of recombinant human erythropoietin (EPO)in CHO cells was explored. A T-flask culture in an incubator without CO(2) addition showed a slow cell growth initially followed by the cessation of growth, while other cultures incubated under 0.5-5% CO(2) concentrations grew normally at the same rate during the entire period of cultivation. Interestingly, the production of EPO in the culture incubated under no CO(2) supply was highest among the tested cultures. The cell specific secretion rate of EPO (q(EPO)) of the culture under no CO(2) supply was about 3 times higher than that of the culture under 5% CO(2) supply. Western blot analysis and in vivo bioassay of EPO showed no apparent changes in EPO quality between the two cases of different CO(2) environments (air vs. 5% CO(2)), suggesting robust glycosylation of EPO by CHO cells even under very reduced CO(2) environment. Various combinations of the two extreme cases, with 5% CO(2) supply (suitable for cell growth) and no CO(2) addition (better for EPO production), were made in order to maximize the volumetric productivity of EPO secretion (P(V)) in CHO cells. The P(V) of the cultures programmed with initial incubation under 5% CO(2) followed by no CO(2) supply was about 2 times superior to that of the culture incubated only under no CO(2) supply. The P(V) of the culture under no CO(2) supply was slightly lower than that of culture grown under 5% CO(2). However, the q(EPO) of the no CO(2) supply case was more than 5 times higher than that of the culture under 5% CO(2) supply. In conclusion, we have demonstrated that a simple programming of CO(2) supply to an incubator can enhance the production of EPO in CHO cells remarkably, without any apparent change of the EPO quality. PMID:19002908

  10. [Correction of anemia in chronic kidney failure with lyophilized recombinant human erythropoietin using a subcutaneous approach].

    PubMed

    Amar, M; Cendoroglo Neto, M; Canziani, M E; Nadaletto, M A; Ajzen, H; Draibe, S A

    1994-01-01

    The recombinant human erythropoietin (rHuEPO) by subcutaneous route has been considered the drug of choice for the correction of the anemia of chronic renal patients. PURPOSE--To evaluate the efficacy of a new preparation of rHuEPO in the correction of the anemia of chronic renal patients maintained by haemodialysis, exclusively administered by subcutaneous route, studying the adverse effects and searching for predictive factors for the response to this medication. METHODS--Twelve patients in regular haemodialysis were treated with freeze-dried rHuEPO by subcutaneous route during 18 months with initial doses of 20U/kg/dialysis. They were submitted to a careful clinical and laboratory monitoring for all this study. RESULTS--Eleven patients ended the study reaching the target hematocrit (Htc) of 30% and keeping it during the whole period of the study. The mean correction and maintenance doses of rHuEPO were 65U/kg/dialysis and 51U/kg/dialysis respectively. At the 12th week of the study a significative increase of Htc (18.4 +/- 3.5% vs. 25.4% +/- 3.8%, p < 0.05) was demonstrated. An increase of the erythrocytes and hemoglobin was concomitantly observed. Leucocytes and platelets increased significantly from the 24th week and kept steadily until the end of study. Just potassium increased in the biochemistry analysis of the patients at the 4th and the 12th week of the study returning to the basal values at the 24th week. The evolution of the iron metabolism parameters demonstrated an intermitent and statistically significant decrease of transferrin saturation at the 1st, 12th and 24th week, returning to the basal levels at the end of study. The serum ferritin did not change (582.7 +/- 700, 9ng/mL vs. 700.0 +/- 651, 6ng/nL). The weight and the blood pressure did not change either, although 2 normotensive patients became hypertensive and 2 others with controlled hypertension needed drug rearrange for blood pressure control (36%). A patient had a seizure episode with a full

  11. Anti-erythropoietin receptor antibodies in systemic lupus erythematosus patients with anemia.

    PubMed

    Luo, X-Y; Yang, M-H; Peng, Ping; Wu, L-J; Liu, Q-S; Chen, L; Tang, Z; Liu, N-T; Zeng, X-F; Liu, Y; Yuan, G-H

    2013-02-01

    Anemia is a common hematologic abnormality in systemic lupus erythematosus (SLE). An inadequate erythropoietin (EPO) response in SLE patients with anemia has been described that may be due to the presence of antibodies to EPO in SLE patients. However, whether anemia in patients with SLE is related to antibodies to EPO receptor (EPOR) has not yet been investigated. We enlisted 169 consecutive patients with SLE and 45 normal individuals to investigate the existence and importance of circulating autoantibodies to EPOR in sera from patients with SLE. In all patients with SLE, the disease activity was evaluated by using the SLE disease activity index SLEDAI. Anti-EPOR antibodies were detected by using an enzyme-linked immunosorbent assay (ELISA). A higher frequency of anti-EPOR antibodies was observed in SLE patients than in healthy controls (18.3% vs 2.2%, p = 0.007). Moreover, anti-EPOR antibodies were detected in 22 of 69 (31.9%) SLE patients with anemia and in only nine of 100 (9.0%, p < 0.001) in those without. Furthermore, the patients with anti-EPOR antibodies exhibited more severe anemia and often presented as microcytic anemia (p = 0.001). Finally, anti-EPOR antibodies seemed more likely to occur in patients with rash (p = 0.008), lower levels of C(3) component (p = 0.01), higher titer of anti-dsDNA antibodies (p < 0.001) and higher disease activity scores (p = 0.024). The results of this study suggest that anti-EPOR antibodies might play a vital role in SLE patients developing anemia because of the higher incidence of antibodies to EPOR found in SLE patients with anemia. Thus, there might be clinical value in detecting anti-EPOR antibodies in SLE patients with anemia. Therefore, the pathologic role of the antibodies in inducing anemia needs to be established in future studies.

  12. Recombinant human erythropoietin in very elderly patients with myelodysplastic syndromes: results from a retrospective study.

    PubMed

    Tatarelli, Caterina; Piccioni, Anna Lina; Maurillo, Luca; Naso, Virginia; Battistini, Roberta; D'Andrea, Mariella; Criscuolo, Marianna; Nobile, Carolina; Villivà, Nicoletta; Mancini, Stefano; Neri, Benedetta; Breccia, Massimo; Fenu, Susanna; Buccisano, Francesco; Voso, Maria Teresa; Latagliata, Roberto; Aloe Spiriti, Maria Antonietta

    2014-08-01

    Myelodysplastic syndromes (MDS) are common in elderly patients. Recombinant human erythro-poietin (rHuEPO) has been widely used to treat anemia in lower risk MDS patients, but few data are known about rHuEPO treatment in the very elderly patient group. In order to investigate the role of rHuEPO treatment in terms of response, overall survival (OS), and toxicity in a very elderly MDS patient group, 93 MDS patients treated with rHuEPO when aged ≥80 years were selected among MDS cases enrolled in a retrospective multicenter study by the cooperative group Gruppo Romano Mielodisplasie (GROM) from Jan 2002 to Dec 2010. At baseline, median age was 82.7 (range 80-99.1) with a median hemoglobin (Hb) level of 9 g/dl (range 6-10.8). The initial dose of rHuEPO was standard (epoetin alpha 40,000 IU/week or epoetin beta 30,000 IU/week) in 59 (63.4 %) patients or high in 34 (36.6 %) (epoetin alpha 80,000 IU/week) patients. We observed an erythroid response (ER) in 59 (63.4 %) patients. No thrombotic event was reported. Independent predictive factors for ER were low transfusion requirement before treatment (p = 0.004), ferritin <200 ng/ml (p = 0.017), Hb >8 g/dl (p = 0.034), and a high-dose rHuEPO treatment (p = 0.032). Median OS from rHuEPO start was 49.3 months (95 % CI 27.5-68.4) in responders versus 30.6 months (95 % CI 7.3-53.8) in resistant patients (p = 0.185). In conclusion, rHuEPO treatment is safe and effective also in the very elderly MDS patients. However, further larger studies are warranted to evaluate if EPO treatment could be worthwhile in terms of quality of life and cost-efficacy in very old patients. PMID:24647684

  13. Mixed-effects modelling of the interspecies pharmacokinetic scaling of pegylated human erythropoietin.

    PubMed

    Jolling, Koen; Perez Ruixo, Juan Jose; Hemeryck, Alex; Vermeulen, An; Greway, Tony

    2005-04-01

    -EPO in humans suggest a less frequent dosing regimen relative to erythropoietin and darbepoetin, potentially leading to a simplification of anemia management.

  14. Function of caspases in regulating apoptosis caused by erythropoietin deprivation in erythroid progenitors.

    PubMed

    Gregoli, P A; Bondurant, M C

    1999-02-01

    Erythropoietin (EP) is required by late stage erythroid progenitor cells to prevent apoptosis. In a previous study (Gregoli and Bondurant, 1997, Blood 90:630-640), it was shown that rapid proteolytic conversion of procaspase 3 to the fully activated enzyme occurred when erythroblasts were deprived of EP for as little as 2 h. In the present study, protein and mRNA analyses of erythroblasts indicated the presence of the proenzyme precursors of caspases 1, 2, 3, 5, 6, 7, 8, and 9. The effects of various caspase inhibitors on caspase 3 processing and on apoptosis were examined. These inhibitors were benzyloxycarbonyl (z-) and fluoromethyl-ketone (FMK) derivatives of peptides that serve as substrates for selected caspases. z-VAD-FMK, t-butoxycarbonyl-aspartate-FMK (Boc-D-FMK), and z-IETD-FMK blocked the initial cleavage of procaspase 3, while z-DEVD-FMK, z-VEID-FMK, and z-VDVAD-FMK did not block the initial cleavage but had some effect on blocking apoptosis. The peptide inhibitor z-FA-FMK, which inhibits cathepsins B and L but is not known to inhibit caspases, altered caspase 3 processing to a final 19 kDa large subunit that appeared to retain enzymatic activity. The action of z-FA-FMK in preventing the usual conversion to a 1 7 kDa subunit suggests the possibility that a noncaspase protease may be involved in caspase 3 processing. Studies with the peptide inhibitors and EP were done to determine the short- and long-term effectiveness of the caspase inhibitors in protecting EP-deprived cells from apoptosis. Although several of the inhibitors were effective, z-IETD-FMK was studied most extensively because of its specificity for enzymes which cleave procaspase 3 at aspartate 175 (IETD175). Large percentages of EP-deprived erythroblasts treated with z-IETD-FMK appeared morphologically normal and negative by a DNA strand breakage (TUNEL) assay at 24 h (75%) compared to EP-deprived controls (10%) which were not treated with inhibitor. However, inhibitor-treated erythroid

  15. Intervention With an Erythropoietin-Derived Peptide Protects Against Neuroglial and Vascular Degeneration During Diabetic Retinopathy

    PubMed Central

    McVicar, Carmel M.; Hamilton, Ross; Colhoun, Liza M.; Gardiner, Tom A.; Brines, Michael; Cerami, Anthony; Stitt, Alan W.

    2011-01-01

    OBJECTIVE 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. RESEARCH DESIGN AND METHODS 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). RESULTS 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. CONCLUSIONS 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

  16. Comparison of two recombinant erythropoietin formulations in patients with anemia due to end-stage renal disease on hemodialysis: A parallel, randomized, double blind study

    PubMed Central

    Pérez-Oliva, Jorge F; Casanova-González, Martha; García-García, Idrian; Porrero-Martín, Pedro J; Valenzuela-Silva, Carmen M; Hernández-Montero, Tairí; Lagarde-Ampudia, Marcia; Casanova-Kutsareva, Yuri; Ávila-Albuerne, Yisel; Vargas-Batista, Alicia; Bobillo-López, Hailen; Herrera-Valdés, Raúl; López-Saura, Pedro A

    2005-01-01

    Background Recombinant human erythropoietin (EPO) is used for the treatment of last stage renal anemia. A new EPO preparation was obtained in Cuba in order to make this treatment fully nationally available. The aim of this study was to compare the pharmacokinetic, pharmacodynamic and safety properties of two recombinant EPO formulations in patients with anemia due to end-stage renal disease on hemodialysis. Methods A parallel, randomized, double blind study was performed. A single 100 IU/Kg EPO dose was administered subcutaneously. Heberitro (Heber Biotec, Havana, formulation A), a newly developed product and Eprex (CILAG AG, Switzerland, formulation B), as reference treatment were compared. Thirty-four patients with anemia due to end-stage renal disease on hemodialysis were included. Patients had not received EPO previously. Serum EPO level was measured by enzyme immunoassay (EIA) during 120 hours after administration. Clinical and laboratory variables were determined as pharmacodynamic and safety criteria until 216 hours. Results Both groups of patients were similar regarding all demographic and baseline characteristics. EPO kinetics profiles were similar for both formulations; the pharmacokinetic parameters were very close (i.e., AUC: 4667 vs. 4918 mIU.h/mL; Cmax: 119.1 vs. 119.7 mIU/mL; Tmax: 13.9 vs. 18.1 h; half-life, 20.0 vs. 22.5 h for formulations A and B, respectively). The 90% confidence intervals for the ratio between both products regarding these metrics were close to the 0.8 – 1.25 range, considered necessary for bioequivalence. Differences did not reach 20% in any case and were not determined by a formulation effect, but probably by a patients' variability effect. Concerning pharmacodynamic features, a high similitude in reticulocyte counts increments until 216 hours and the percentage decrease in serum iron until 120 hours was observed. There were no differences between formulations regarding the adverse events and their intensity. The more

  17. Lentiviral vector platform for improved erythropoietin expression concomitant with shRNA mediated host cell elastase down regulation.

    PubMed

    Dhamne, Hemant; Chande, Ajit G; Mukhopadhyaya, Robin

    2014-01-01

    Lentiviral vector (LV) mediated gene transfer holds great promise to develop stable cell lines for sustained transgene expression providing a valuable alternative to the conventional plasmid transfection based recombinant protein production methods. We report here making a third generation HIV-2 derived LV containing erythropoietin (EPO) gene expression cassette to generate a stable HEK293 cell line secreting EPO constitutively. A high producer cell clone was obtained by limiting dilution and was adapted to serum free medium. The suspension adapted cell clone stably produced milligram per liter quantities of EPO. Subsequent host metabolic engineering using lentiviral RNAi targeted to block an endogenous candidate protease elastase, identified through an in silico approach, resulted in appreciable augmentation of EPO expression above the original level. This study of LV based improved glycoprotein expression with host cell metabolic engineering for stable production of protein therapeutics thus exemplifies the versatility of LV and is of significant future biopharmaceutical importance.

  18. Erythropoietin attenuates Alzheimer-like memory impairments and pathological changes induced by amyloid β42 in mice.

    PubMed

    Li, Yi-Pei; Yang, Guo-Jun; Jin, Li; Yang, Hong-Mei; Chen, Jie; Chai, Gao-Shang; Wang, Li

    2015-08-27

    Amyloid beta (Aβ) is a key molecule in the neurodegenerative progression of Alzheimer׳s disease (AD). It is critical to develop a treatment that can arrest the Aβ-induced pathologic progression of AD. Erythropoietin (EPO) has various protective effects in the nervous system. However, the effect of EPO on Aβ-induced Alzheimer-like cognitive deficits and pathological changes remains unclear. In the present study, we observed that the treatment of mice with EPO (1000 IU/kg) attenuated Aβ42-induced cognitive deficits and tau hyperphosphorylation at multiple AD-related sites through the regulation of glycogen synthase kinase-3β (GSK-3β). We also observed that EPO attenuated the Aβ42-induced mitochondrial dysfunction and apoptosis in brain. These results indicate a potential role for EPO in AD therapy.

  19. Lentiviral vector platform for improved erythropoietin expression concomitant with shRNA mediated host cell elastase down regulation.

    PubMed

    Dhamne, Hemant; Chande, Ajit G; Mukhopadhyaya, Robin

    2014-01-01

    Lentiviral vector (LV) mediated gene transfer holds great promise to develop stable cell lines for sustained transgene expression providing a valuable alternative to the conventional plasmid transfection based recombinant protein production methods. We report here making a third generation HIV-2 derived LV containing erythropoietin (EPO) gene expression cassette to generate a stable HEK293 cell line secreting EPO constitutively. A high producer cell clone was obtained by limiting dilution and was adapted to serum free medium. The suspension adapted cell clone stably produced milligram per liter quantities of EPO. Subsequent host metabolic engineering using lentiviral RNAi targeted to block an endogenous candidate protease elastase, identified through an in silico approach, resulted in appreciable augmentation of EPO expression above the original level. This study of LV based improved glycoprotein expression with host cell metabolic engineering for stable production of protein therapeutics thus exemplifies the versatility of LV and is of significant future biopharmaceutical importance. PMID:24325878

  20. Plasminogen activator inhibitor-1 fused with erythropoietin (EPO) mimetic peptide (EMP) enhances the EPO activity of EMP.

    PubMed

    Kuai, L; Wu, C; Qiu, Q; Zhang, J; Zhou, A; Wang, S; Zhang, H; Song, Q; Liao, S; Han, Y; Liu, J; Ma, Z

    2000-08-01

    Erythropoietin (EPO) mimetic peptide (EMP) encoding sequence was inserted into the gene of plasminogen activator inhibitor-1 (PAI-1) between Ala348 and Pro349 (P2'-P3'), generating a novel gene, PAI-1/EMP (PMP). This was cloned into pET32a expression vector, fused with TrxA peptide in the vector, and a 63-kDa protein was expressed in inclusion bodies with an expression level >50%. The TrxA/PMP protein was purified by Ni-NTA-agarose metal-ligand affinity chromatography to a purity >90%, showing a single, silver-stained band on SDS-PAGE. Using a reticulocyte counting assay, the EPO activity of PMP was determined to be 5,000 IU/mg, 2,500-fold that of EMP.

  1. Modulation of cellular stress response via the erythropoietin/CD131 heteroreceptor complex in mouse mesenchymal-derived cells.

    PubMed

    Bohr, Stefan; Patel, Suraj J; Vasko, Radovan; Shen, Keyue; Iracheta-Vellve, Arvin; Lee, Jungwoo; Bale, Shyam Sundhar; Chakraborty, Nilay; Brines, Michael; Cerami, Anthony; Berthiaume, Francois; Yarmush, Martin L

    2015-02-01

    Tissue-protective properties of erythropoietin (EPO) have let to the discovery of an alternative EPO signaling via an EPO-R/CD131 receptor complex which can now be specifically targeted through pharmaceutically designed short sequence peptides such as ARA290. However, little is still known about specific functions of alternative EPO signaling in defined cell populations. In this study, we investigated effects of signaling through EPO-R/CD131 complex on cellular stress responses and pro-inflammatory activation in different mesenchymal-derived phenotypes. We show that anti-apoptotic, anti-inflammatory effects of ARA290 and EPO coincide with the externalization of CD131 receptor component as an immediate response to cellular stress. In addition, alternative EPO signaling strongly modulated transcriptional, translational, or metabolic responses after stressor removal. Specifically, we saw that ARA290 was able to overcome a TNFα-mediated inhibition of transcription factor activation related to cell stress responses, most notably of serum response factor (SRF), heat shock transcription factor protein 1 (HSF1), and activator protein 1 (AP1). We conclude that alternative EPO signaling acts as a modulator of pro-inflammatory signaling pathways and likely plays a role in restoring tissue homeostasis. Key message: Erythropoietin (EPO) triggers an alternative pathway via heteroreceptor EPO/CD131. ARA290 peptide specifically binds EPO/CD131 but not the canonical EPO/EPO receptor. Oxidative stress and inflammation promote cell surface expression of CD131. ARA290 prevents tumor necrosis factor-mediated inhibition of stress-related genes. Alternative EPO signaling modulates inflammation and promotes tissue homeostasis.

  2. Efficient preparation and PEGylation of recombinant human non-glycosylated erythropoietin expressed as inclusion body in E. coli.

    PubMed

    Wang, Yin-Jue; Liu, Yong-Dong; Chen, Jing; Hao, Su-Juan; Hu, Tao; Ma, Guang-Hui; Su, Zhi-Guo

    2010-02-15

    Recombinant human erythropoietin produced by mammalian cells contains about 40% carbohydrates which maintain its stability and long residence in body. However, mammalian derived Epo has low yields and high costs of production. In this article, a cost-effective strategy of producing non-glycosylated Epo from Escherichia coli and then PEGylating it to replace the role of sugar chains was investigated. Recombinant human non-glycosylated erythropoietin (rh-ngEpo) was overexpressed as inclusion body in E. coli. As the routine inclusion body washing step resulted in poor protein recovery and purity, a new process scheme of using strong ion-exchange chromatography to purify denatured rh-ngEpo from inclusion body before refolding was developed. The purity of the denatured rh-ngEpo was increased from 59% to over 90%. Rh-ngEpo was then refolded and subsequently purified by one step of weak cation-exchange chromatography to 98% pure. Final protein yield was 129 mg/l, a significant improvement from 49 mg/l obtained via the conventional practice. The in vitro bioactivity of purified rh-ngEpo was comparable with the CHO-expressed Epo and the formation of native secondary structure was also confirmed by CD spectra. Rh-ngEpo was then modified by a 20 kDa methoxy polyethylene glycol (PEG) succinimidyl carbonate. The monoPEGylated protein, which retained 68% bioactivity, had enhanced thermal stability and a remarkably prolonged circulating half-life in rats as compared with that of the unmodified protein. These studies demonstrated the feasibility of PEGylating rh-ngEpo as a promising way for the development of new Epo drugs.

  3. The influence of Erythropoietin on platelet activation, thrombin generation and FVII/active FVII in patients with AMI

    PubMed Central

    2014-01-01

    Background Erythropoietin (Epo) has been shown to improve myocardial function in models of experimental myocardial infarction, but has also been associated with a rise in thromboembolic events. Thus, the aim of this study was to investigate the influence of Epo on platelet activation and coagulation in patients with acute myocardial infarction (AMI). Methods The study was designed as a substudy of the randomised, double-blind, placebo controlled REVIVAL-3 (REgeneration of VItal Myocardium in ST-Segment EleVation MyocardiAL Infarction by Erythropoietin) study that investigated the effects of recombinant human Epo in AMI. Serial venous blood samples were collected before and after study medication. Circulating prothrombin fragment F1 + 2, FVII, active FVII, beta thromboglobulin (TG) and P-Selectin were measured before and 60 hours after randomization by immunoassay (n = 94). In a randomly selected subgroup platelet aggregation was measured using whole blood aggregometry (Multiplate Analyzer, n = 45). Results After 5 days an increase in FVII was observed after Epo as compared to placebo (P = 0.02), yet active FVII and prothrombin fragment F1 + 2 remained unchanged. Moreover, no statistically significant differences in circulating TG or P-selectin were observed between the groups. As an expected response to peri-interventional therapy with clopidogrel and aspirin, platelet aggregation after stimulation with ADP, TRAP, ASPI or collagen decreased 12 hours and 2 days after PCI. However, no difference between the Epo and the placebo group was observed. Conclusion After treatment with Epo in patients with AMI a slight increase in circulating FVII after Epo was not associated with an increase in active FVII, prothrombin fragment F1 + 2, TG or P-selectin. Moreover, platelet aggregation was not altered after treatment with Epo as compared to placebo. Trial registration ClinicalTrials.gov Identifier: NCT01761435 PMID:25228850

  4. Influence of recombinant human erythropoietin treatment on pulmonary O2 uptake kinetics during exercise in humans

    PubMed Central

    Wilkerson, Daryl P; Rittweger, Jörn; Berger, Nicolas JA; Naish, Patrick F; Jones, Andrew M

    2005-01-01

    We hypothesized that 4 weeks of recombinant human erythropoietin (RhEPO) treatment would result in a significant increase in haemoglobin concentration ([Hb]) and arterial blood O2-carrying capacity and that this would (1) increase peak pulmonary oxygen uptake V˙O2 during ramp incremental exercise, and (2) speed V˙O2 kinetics during ‘severe’-, but not ‘moderate’- or ‘heavy’-intensity, step exercise. Fifteen subjects (mean ±s.d. age 25 ± 4 years) were randomly assigned to either an experimental group which received a weekly subcutaneous injection of RhEPO (150 IU kg−1; n = 8), or a control group (CON) which received a weekly subcutaneous injection of sterile saline (10 ml; n = 7) as a placebo, for four weeks. The subjects and the principal researchers were both blind with respect to the group assignment. Before and after the intervention period, all subjects completed a ramp test for determination of the gas exchange threshold (GET) and V˙O2,peak, and a number of identical ‘step’ transitions from ‘unloaded’ cycling to work rates requiring 80% GET (moderate), 70% of the difference between the GET and V˙O2,peak (heavy), and 105% V˙O2,peak (severe) as determined from the initial ramp test. Pulmonary gas exchange was measured breath-by-breath. There were no significant differences between the RhEPO and CON groups for any of the measurements of interest ([Hb], V˙O2,peak V˙O2 kinetics) before the intervention. Four weeks of RhEPO treatment resulted in a 7% increase both in [Hb] (from 15.8 ± 1.0 to 16.9 ± 0.7 g dl−1; P < 0.01) and V˙O2,peak (from 47.5 ± 4.2 to 50.8 ± 10.7 ml kg−1·min−1; P < 0.05), with no significant change in CON. RhEPO had no significant effect on V˙O2 kinetics for moderate (Phase II time constant, from 28 ± 8 to 28 ± 7 s), heavy (from 37 ± 12 to 35 ± 11 s), or severe (from 33 ± 15 to 35 ± 15 s) step exercise. Our results indicate that enhancing blood O2-carrying capacity and thus the potential for muscle

  5. The phenomenon of "pre-ischaemic conditioning" in the brain only partly involves the NMDA receptor: a magnetic resonance study.

    PubMed

    Prior, Malcolm; Thatcher, Nicola; Morris, Peter; Reese, Torsten; Bachelard, Herman

    2005-10-01

    We have investigated in more detail our previous observations on a form of ischaemic pre-conditioning "metabolic adaptation", i.e.--that sequential metabolic insults (hypoxia followed 40 min later by combined hypoxia + hypoglycaemia, or vice versa) are less injurious (monitored by increased [Ca2+]i and decreased PCr) than the immediate combined insult. We have now observed that the "adaptation" occurs between 10 and 20 min. Pre-treatment of the tissues with 10 microM-MK801 showed that it had no effect on the increase in [Ca2+]i caused by the sequential insult and only partially blocked the increase observed by exposure to the immediate combined insult. Exposure to both the delayed and immediate combined insults with low extracellular Ca2+ resulted in a two-fold increase in [Ca2+]i, similar to the increase observed with normal extracellular Ca2+ in the presence of MK801. The results are discussed in terms of the possible origins of the increases in [Ca2+]i.

  6. Fabrication of an on-line enzyme micro-reactor coupled to liquid chromatography-tandem mass spectrometry for the digestion of recombinant human erythropoietin.

    PubMed

    Foo, Hsiao Ching; Smith, Norman W; Stanley, Shawn M R

    2015-04-01

    Our aim was to develop a fast and efficient on-line method using micro-reactors for the digestion and deglycosylation of recombinant human erythropoietin extracted from equine plasma. The trypsin digestion micro reactors were fabricated using fused silica capillaries with either a dextran-modified coating or a porous monolith that was able to immobilise the enzyme. These were both found to be reasonably robust and durable, with the trypsin immobilised on dextran-modified fused silica capillaries offering better reproducibility than the micro-reactor based upon covalent attachment of this enzyme to the polymer. It is also evident that the enzyme attached micro reactors produced some tryptic peptides in a greater yield than in-solution digestion. A peptide-N-glycosidase F reactor was also fabricated and, when coupled with the trypsin reactor, the deaminated peptides T5 DAM and T9 DAM from recombinant human erythropoietin could also be detected by LC-ESI-MS/MS analysis. These results were better than those achieved using off-line digestion plus deglycosylation reactions and the analysis required far less time and effort to complete. The use of this on-line approach improved the sensitivity, efficiency and speed of our confirmation methodology that is based upon detecting the unique peptide segments of recombinant human erythropoietin that has been affinity extracted from positive equine plasma samples.

  7. PI3K and MEK1/2 molecular pathways are involved in the erythropoietin-mediated regulation of the central respiratory command.

    PubMed

    Caravagna, Céline; Soliz, Jorge

    2015-01-15

    Erythropoietin stimulation modulates the central respiratory command in newborn mice. Specifically, the central respiratory depression induced by hypoxia is attenuated by acute (1h) or abolished by chronic erythropoietin stimulation. However, the underlying mechanisms remain unknown. As MEK and PI3K pathways are commonly involved in Epo-mediated effects of neuroprotection and erythropoiesis, we investigated here the implication of PI3K and MEK1/2 in the Epo-mediated regulation of the central respiratory command. To this end, in vitro brainstem-spinal cord preparations from 3 days old transgenic (Tg21; constitutively overexpressing erythropoietin in the brain specifically) and control mice were used. Our results show that blockade of PI3K or MEK1/2 stimulates normoxic bursts frequency in Tg21 preparations and abolish hypoxia-induced frequency depression in control preparations. These results show that MEK1/2 and PI3K pathways are involved in the Epo-mediated regulation of the central respiratory command. Moreover, this is the first demonstration that MEK1/2 and PI3K are involved in the brainstem central respiratory command. PMID:25462838

  8. Post-treatment Effects of Erythropoietin and Nordihydroguaiaretic Acid on Recovery from Cisplatin-induced Acute Renal Failure in the Rat

    PubMed Central

    Lee, Dong Won; Lee, Soo Bong; Song, Sang Heon; Seong, Eun Young; Yang, Byeong Yun; Lee, Min Young; Sol, Mee Young

    2009-01-01

    5-Lipoxygenase inhibitor and human recombinant erythropoietin might accelerate renal recovery in cisplatin-induced acute renal failure rats. Male Sprague-Dawley rats were randomized into four groups: 1) normal controls; 2) Cisplatin group-cisplatin induced acute renal failure (ARF) plus vehicle treatment; 3) Cisplatin+nordihydroguaiaretic acid (NDGA) group-cisplatin induced ARF plus 5-lipoxygenase inhibitor treatment; 4) Cisplatin+erythropoietin (EPO) group-cisplatin induced ARF plus erythropoietin treatment. On day 10 (after 7 daily injections of NDGA or EPO), urea nitrogen and serum Cr concentrations were significantly lower in the Cisplatin+NDGA and Cisplatin+EPO groups than in the Cisplatin group, and 24 hr urine Cr clearances were significantly higher in the Cisplatin+EPO group than in the Cisplatin group. Semi-quantitative assessments of histological lesions did not produce any significant differences between the three treatment groups. Numbers of PCNA(+) cells were significantly higher in Cisplatin, Cisplatin+NDGA, and Cisplatin+EPO groups than in normal controls. Those PCNA(+) cells were significantly increased in Cisplatin+NDGA group. These results suggest that EPO and also NDGA accelerate renal function recovery by stimulating tubular epithelial cell regeneration. PMID:19194548

  9. Transportation Security Administration

    MedlinePlus

    ... content Official website of the Department of Homeland Security Transportation Security Administration A - Z Index What Can I Bring? Search form Apples Main menu Administrator Travel Security Screening Special Procedures TSA Pre✓® Passenger Support Travel ...

  10. NASA, NOAA administrators nominated

    NASA Astrophysics Data System (ADS)

    Richman, Barbara T.

    President Ronald Reagan recently said he intended to nominate James Montgomery Beggs as NASA Administrator and John V. Byrne as NOAA Administrator. These two positions are key scientific posts that have been vacant since the start of the Reagan administration on January 20. The President also said he intends to nominate Hans Mark as NASA Deputy Administrator. At press time, Reagan had not designated his nominee for the director of the Office of Science and Technology Policy.

  11. A Philosophy of Administration.

    ERIC Educational Resources Information Center

    Bruening, William H.

    Justification is given for paying relatively large salaries to college administrators, specifically the president or chancellor and the chief academic officer. Three administrative task areas are discussed as criteria: management, administration per se, and leadership. It is contended that only leadership can be used as a criterion for…

  12. Identification of Cell Type-Specific Differences in Erythropoietin Receptor Signaling in Primary Erythroid and Lung Cancer Cells

    PubMed Central

    Salopiata, Florian; Depner, Sofia; Wäsch, Marvin; Böhm, Martin E.; Mücke, Oliver; Plass, Christoph; Lehmann, Wolf D.; Kreutz, Clemens; Timmer, Jens; Klingmüller, Ursula

    2016-01-01

    Lung cancer, with its most prevalent form non-small-cell lung carcinoma (NSCLC), is one of the leading causes of cancer-related deaths worldwide, and is commonly treated with chemotherapeutic drugs such as cisplatin. Lung cancer patients frequently suffer from chemotherapy-induced anemia, which can be treated with erythropoietin (EPO). However, studies have indicated that EPO not only promotes erythropoiesis in hematopoietic cells, but may also enhance survival of NSCLC cells. Here, we verified that the NSCLC cell line H838 expresses functional erythropoietin receptors (EPOR) and that treatment with EPO reduces cisplatin-induced apoptosis. To pinpoint differences in EPO-induced survival signaling in erythroid progenitor cells (CFU-E, colony forming unit-erythroid) and H838 cells, we combined mathematical modeling with a method for feature selection, the L1 regularization. Utilizing an example model and simulated data, we demonstrated that this approach enables the accurate identification and quantification of cell type-specific parameters. We applied our strategy to quantitative time-resolved data of EPO-induced JAK/STAT signaling generated by quantitative immunoblotting, mass spectrometry and quantitative real-time PCR (qRT-PCR) in CFU-E and H838 cells as well as H838 cells overexpressing human EPOR (H838-HA-hEPOR). The established parsimonious mathematical model was able to simultaneously describe the data sets of CFU-E, H838 and H838-HA-hEPOR cells. Seven cell type-specific parameters were identified that included for example parameters for nuclear translocation of STAT5 and target gene induction. Cell type-specific differences in target gene induction were experimentally validated by qRT-PCR experiments. The systematic identification of pathway differences and sensitivities of EPOR signaling in CFU-E and H838 cells revealed potential targets for intervention to selectively inhibit EPO-induced signaling in the tumor cells but leave the responses in erythroid

  13. Memory Improvement in the AβPP/PS1 Mouse Model of Familial Alzheimer’s Disease Induced by Carbamylated-Erythropoietin is Accompanied by Modulation of Synaptic Genes.

    PubMed

    Armand-Ugón, Mercedes; Aso, Ester; Moreno, Jesús; Riera-Codina, Miquel; Sánchez, Alex; Vegas, Esteban; Ferrer, Isidre

    2015-01-01

    Neuroprotection of erythropoietin (EPO) following long-term administration is hampered by the associated undesirable effects on hematopoiesis and body weight. For this reason, we tested carbamylated-EPO (CEPO), which has no effect on erythropoiesis, and compared it with EPO in the AβPP/PS1 mouse model of familial Alzheimer’s disease. Groups of 5-month old wild type (WT) and transgenic mice received chronic treatment consisting of CEPO (2,500 or 5,000 UI/kg) or EPO (2,500 U I/kg) 3 days/week for 4 weeks. Memory at the end of treatment was assessed with the object recognition test. Microarray analysis and quantitative-PCR were used for gene expression studies. No alterations in erythropoiesis were observed in CEPO-treated WT and AβPP/PS1 transgenic mice. EPO and CEPO improved memory in AβPP/PS1 animals. However, only EPO decreased amyloid-β (Aβ)plaque burden and soluble Aβ(40). Microarray analysis of gene expression revealed a limited number of common genes modulated by EPO and CEPO. CEPO but not EPO significantly increased gene expression of dopamine receptors 1 and 2, and adenosine receptor 2a, and significantly down-regulated adrenergic receptor 1D and gastrin releasing peptide. CEPO treatment resulted in higher protein levels of dopamine receptors 1 and 2 in WT and AβPP/PS1 animals, whereas the adenosine receptor 2a was reduced in WT animals. The present results suggest that the improved behavior observed in AβPP/PS1 transgenic mice after CEPO treatment may be mediated, at least in part, by the observed modulation of the expression of molecules involved in neurotransmission. PMID:25790933

  14. Erythropoietin-mediated expression of placenta growth factor is regulated via activation of hypoxia-inducible factor-1α and post-transcriptionally by miR-214 in sickle cell disease.

    PubMed

    Gonsalves, Caryn S; Li, Chen; Mpollo, Marthe-Sandrine Eiymo Mwa; Pullarkat, Vinod; Malik, Punam; Tahara, Stanley M; Kalra, Vijay K

    2015-06-15

    Placental growth factor (PlGF) plays an important role in various pathological conditions and diseases such as inflammation, cancer, atherosclerosis and sickle cell disease (SCD). Abnormally high PlGF levels in SCD patients are associated with increased inflammation and pulmonary hypertension (PHT) and reactive airway disease; however, the transcriptional and post-transcriptional mechanisms regulating PlGF expression are not well defined. Herein, we show that treatment of human erythroid cells and colony forming units with erythropoietin (EPO) increased PlGF expression. Our studies showed EPO-mediated activation of HIF-1α led to subsequent binding of HIF-1α to hypoxia response elements (HREs) within the PlGF promoter, as demonstrated by luciferase transcription reporter assays and ChIP analysis of the endogenous gene. Additionally, we showed miR-214 post-transcriptionally regulated the expression of PlGF as demonstrated by luciferase reporter assays using wild-type (wt) and mutant PlGF-3'-UTR constructs. Furthermore, synthesis of miR-214, located in an intron of DNM3 (dynamin 3), was transcriptionally regulated by transcription factors, peroxisome proliferator-activated receptor-α (PPARα) and hypoxia-inducible factor-1α (HIF-1α). These results were corroborated in vivo wherein plasma from SCD patients and lung tissues from sickle mice showed an inverse correlation between PlGF and miR-214 levels. Finally, we observed that miR-214 expression could be induced by fenofibrate, a Food and Drug Administration (FDA) approved PPARα agonist, thus revealing a potential therapeutic approach for reduction in PlGF levels by increasing miR-214 transcription. This strategy has potential clinical implications for several pathological conditions including SCD.

  15. Increased bone formation in a rabbit long-bone defect model after single local and single systemic application of erythropoietin.

    PubMed

    Omlor, Georg W; Kleinschmidt, Kerstin; Gantz, Simone; Speicher, Anja; Guehring, Thorsten; Richter, Wiltrud

    2016-08-01

    Background and purpose - Delayed bone healing with non-union is a common problem. Further options to increase bone healing together with surgery are needed. We therefore evaluated a 1-dose single application of erythropoietin (EPO), applied either locally to the defect or systemically during surgery, in a critical-size rabbit long-bone defect. Material and methods - 19 New Zealand White rabbits received a 15-mm defect in the radius diaphysis. An absorbable gelatin sponge was soaked with saline (control group and systemic treatment group) or EPO (local treatment group) and implanted into the gap. The systemic treatment group received EPO subcutaneously. In vivo micro-CT analysis was performed 4, 8, and 12 weeks postoperatively. Vascularization was evaluated histologically. Results - Semiquantitative histomorphometric and radiological evaluation showed increased bone formation (2.3- to 2.5-fold) in both treatment groups after 12 weeks compared to the controls. Quantitative determination of bone volume and tissue volume showed superior bone healing after EPO treatment at all follow-up time points, with the highest values after 12 weeks in locally treated animals (3.0- to 3.4-fold). More vascularization was found in both EPO treatment groups. Interpretation - Initial single dosing with EPO was sufficient to increase bone healing substantially after 12 weeks of follow-up. Local application inside the defect was most effective, and it can be administered directly during surgery. Apart from effects on ossification, systemic and local EPO treatment leads to increased callus vascularization. PMID:27348783

  16. A systematic scan of interactions with tyrosine motifs in the erythropoietin receptor using a mammalian 2-hybrid approach.

    PubMed

    Montoye, Tony; Lemmens, Irma; Catteeuw, Dominiek; Eyckerman, Sven; Tavernier, Jan

    2005-06-01

    Signaling via the erythropoietin receptor (EpoR) depends on the interaction of several proteins with phosphorylated tyrosine-containing motifs in its cytosolic domain. Detailed mapping of these interactions is required for an accurate insight into Epo signaling. We recently developed a mammalian protein-protein interaction trap (MAPPIT), a cytokine receptor-based 2-hybrid method that operates in intact Hek293-T mammalian cells. As baits, we used intracellular segments of the EpoR containing 1 or 2 tyrosines. Several known signaling molecules, including cytokine-inducible SH2-containing protein (CIS), suppressor of cytokine signaling-2 (SOCS2), phosphatidylinositol 3'-kinase (PI3-K), phospholipase C-gamma (PLC-gamma), and signal transducer and activator of transcription 5 (STAT5) were used as prey. We also extended the MAPPIT method to enable interaction analysis with wild-type EpoR. In this relay MAPPIT approach, instead of using isolated EpoR fragments as bait, we used the full-length EpoR itself as a "receptor bait." Finally, we introduced MAPPIT in the erythroleukemic TF-1 cell line, which is a more natural setting of the EpoR. With these strategies several known interactions with the EpoR were analyzed and evidence for new interactions was obtained. PMID:15644415

  17. Alpha4beta1 integrin and erythropoietin mediate temporally distinct steps in erythropoiesis: integrins in red cell development.

    PubMed

    Eshghi, Shawdee; Vogelezang, Mariette G; Hynes, Richard O; Griffith, Linda G; Lodish, Harvey F

    2007-06-01

    Erythropoietin (Epo) is essential for the terminal proliferation and differentiation of erythroid progenitor cells. Fibronectin is an important part of the erythroid niche, but its precise role in erythropoiesis is unknown. By culturing fetal liver erythroid progenitors, we show that fibronectin and Epo regulate erythroid proliferation in temporally distinct steps: an early Epo-dependent phase is followed by a fibronectin-dependent phase. In each phase, Epo and fibronectin promote expansion by preventing apoptosis partly through bcl-xL. We show that alpha(4), alpha(5), and beta(1) are the principal integrins expressed on erythroid progenitors; their down-regulation during erythropoiesis parallels the loss of cell adhesion to fibronectin. Culturing erythroid progenitors on recombinant fibronectin fragments revealed that only substrates that engage alpha(4)beta(1)-integrin support normal proliferation. Collectively, these data suggest a two-phase model for growth factor and extracellular matrix regulation of erythropoiesis, with an early Epo-dependent, integrin-independent phase followed by an Epo-independent, alpha(4)beta(1)-integrin-dependent phase.

  18. Role of erythropoietin in the angiogenic activity of bone marrow endothelial cells of MGUS and multiple myeloma patients.

    PubMed

    Lamanuzzi, Aurelia; Saltarella, Ilaria; Ferrucci, Arianna; Ria, Roberto; Ruggieri, Simona; Racanelli, Vito; Rao, Luigia; Annese, Tiziana; Nico, Beatrice; Vacca, Angelo; Ribatti, Domenico

    2016-03-22

    Increasing evidences suggest several biological roles for erythropoietin and its receptor (Epo and EpoR), unrelated to erythropoiesis, including angiogenesis. Here, we detected the expression of EpoR in bone marrow-derived endothelial cells from monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM) patients (MGECs and MMECs, respectively) and assessed whether Epo plays a role in MGECs- and MMECs-mediated angiogenesis. We show that EpoR is expressed by both MGECs and MMECs even though at a higher level in the first ones. Both EC types respond to rHuEpo in terms of cell proliferation, whereas other responses, including activation of JAK2/STAT5 and PI3K/Akt pathways, cell migration and capillarogenesis are enhanced by Epo in MGECs, but not in MMECs. In addition, the conditioned media of both Epo-treated cells induce a strong angiogenic response in vivo in the chorioallantoic membrane assay, comparable to that of vascular endothelial growth factor (VEGF). Overall, these data highlight the effect of Epo on MGECs- and MMECs-mediated angiogenesis: MGECs are more responsive to Epo treatment than MMECs, probably because over-angiogenic phenotype of MMECs is already activated by their autocrine/paracrine loops occurring in the "angiogenic switch" from MGUS. PMID:26919105

  19. Expression of constitutively active erythropoietin receptor in pyramidal neurons of cortex and hippocampus boosts higher cognitive functions in mice

    PubMed Central

    2011-01-01

    Background Erythropoietin (EPO) and its receptor (EPOR) are expressed in the developing brain and their transcription is upregulated in adult neurons and glia upon injury or neurodegeneration. We have shown neuroprotective effects and improved cognition in patients with neuropsychiatric diseases treated with EPO. However, the critical EPO targets in brain are unknown, and separation of direct and indirect effects has remained difficult, given the role of EPO in hematopoiesis and brain oxygen supply. Results Here we demonstrate that mice with transgenic expression of a constitutively active EPOR isoform (cEPOR) in pyramidal neurons of cortex and hippocampus exhibit enhancement of spatial learning, cognitive flexibility, social memory, and attentional capacities, accompanied by increased impulsivity. Superior cognitive performance is associated with augmented long-term potentiation of cEPOR expressing neurons in hippocampal slices. Conclusions Active EPOR stimulates neuronal plasticity independent of any hematopoietic effects and in addition to its neuroprotective actions. This property of EPOR signaling should be exploited for defining novel strategies to therapeutically enhance cognitive performance in disease conditions. PMID:21527022

  20. β Common Receptor Mediates Erythropoietin-Conferred Protection on OxLDL-Induced Lipid Accumulation and Inflammation in Macrophages

    PubMed Central

    Lu, Kuo-Yun; Yu, Yuan-Bin; Tsai, Feng-Chuan

    2015-01-01

    Erythropoietin (EPO), the key factor for erythropoiesis, also protects macrophage foam cells from lipid accumulation, yet the definitive mechanisms are not fully understood. β common receptor (βCR) plays a crucial role in the nonhematopoietic effects of EPO. In the current study, we investigated the role of βCR in EPO-mediated protection in macrophages against oxidized low-density lipoprotein- (oxLDL-) induced deregulation of lipid metabolism and inflammation. Here, we show that βCR expression was mainly in foamy macrophages of atherosclerotic aortas from apolipoprotein E-deficient mice. Results of confocal microscopy and immunoprecipitation analyses revealed that βCR was colocalized and interacted with EPO receptor (EPOR) in macrophages. Inhibition of βCR activation by neutralizing antibody or small interfering RNA (siRNA) abolished the EPO-conferred protection in oxLDL-induced lipid accumulation. Furthermore, EPO-promoted cholesterol efflux and upregulation of ATP-binding cassette (ABC) transporters ABCA1 and ABCG1 were prevented by pretreatment with βCR neutralizing antibody or βCR siRNA. Additionally, blockage of βCR abrogated the EPO-conferred anti-inflammatory action on oxLDL-induced production of macrophage inflammatory protein-2. Collectively, our findings suggest that βCR may play an important role in the beneficial effects of EPO against oxLDL-elicited dysfunction of macrophage foam cells. PMID:26101463

  1. Erythropoietin Levels Increase during Cerebral Malaria and Correlate with Heme, Interleukin-10 and Tumor Necrosis Factor-Alpha in India.

    PubMed

    Dalko, Esther; Tchitchek, Nicolas; Pays, Laurent; Herbert, Fabien; Cazenave, Pierre-André; Ravindran, Balachandran; Sharma, Shobhona; Nataf, Serge; Das, Bidyut; Pied, Sylviane

    2016-01-01

    Cerebral malaria (CM) caused by Plasmodium falciparum parasites often leads to the death of infected patients or to persisting neurological sequelae despite anti-parasitic treatments. Erythropoietin (EPO) was recently suggested as a potential adjunctive treatment for CM. However diverging results were obtained in patients from Sub-Saharan countries infected with P. falciparum. In this study, we measured EPO levels in the plasma of well-defined groups of P. falciparum-infected patients, from the state of Odisha in India, with mild malaria (MM), CM, or severe non-CM (NCM). EPO levels were then correlated with biological parameters, including parasite biomass, heme, tumor necrosis factor (TNF)-α, interleukin (IL)-10, interferon gamma-induced protein (IP)-10, and monocyte chemoattractant protein (MCP)-1 plasma concentrations by Spearman's rank and multiple correlation analyses. We found a significant increase in EPO levels with malaria severity degree, and more specifically during fatal CM. In addition, EPO levels were also found correlated positively with heme, TNF-α, IL-10, IP-10 and MCP-1 during CM. We also found a significant multivariate correlation between EPO, TNF-α, IL-10, IP-10 MCP-1 and heme, suggesting an association of EPO with a network of immune factors in CM patients. The contradictory levels of circulating EPO reported in CM patients in India when compared to Africa highlights the need for the optimization of adjunctive treatments according to the targeted population. PMID:27441662

  2. Effect of Recombinant Human Erythropoietin On the Stemness of Bone Marrow-derived Mesenchymal Stem Cells in vitro

    PubMed Central

    Ye, Lihua; Chen, Long; Yu, Qiang; Cheng, Fanjun

    2010-01-01

    The purpose of this study was to investigate the effects of the recombinant human erythropoietin (rhEPO) on proliferative and multi-differentiation potential of the bone marrow-derived mesenchymal stem cells (MSCs). The MSCs were isolated primarily from bone marrow of adult rat and purified at increasing passage. A purified population of MSCs can be obtained about 2 weeks after the initiation of culture. After three passages (P3-MSCs), bone marrow-derived adherent cells were identified, then different concentrations of rhEPO (0.1, 1, 5, 10, 100 U/ml) was added into the Passage-3 cells which had been identified. The expression of the surface markers in adherent cells was detected by the flow cytometry. The mRNA levels of transcription factors OCT4, SOX2, Nanog and TERT were measured by reverse transcription-polymerase chain reaction (RT-PCR). The results showed that CD29 and CD90 were positive in MSCs, but not CD33, CD44 and CD45, and the cells could differentiate into multiple lineages such as osteocytes and adipocytes. The expression of OCT4, SOX2, TERT, Nanog mRNA were up-regulated by the treatment of EPO. The effect of EPO was the most obvious when its concentration was 5U/mL after 12h. we conclude that MSCs can not only perserve characteristics of stem cells but also maintain its multi-lineage differentiation potential after appropriate treatment of EPO. PMID:24855555

  3. Generation and phenotypic analysis of a transgenic line of rabbits secreting active recombinant human erythropoietin in the milk.

    PubMed

    Mikus, Tomás; Poplstein, Martin; Sedláková, Jirina; Landa, Vladimír; Jeníkova, Gabriela; Trefil, Pavel; Lidický, Jan; Malý, Petr

    2004-10-01

    Production of recombinant human erythropoietin (rhEPO) for therapeutic purposes relies on its expression in selected clones of transfected mammalian cells. Alternatively, this glycoprotein can be produced by targeted secretion into the body fluid of transgenic mammals. Here, we report on the generation of a transgenic rabbits producing rhEPO in the lactating mammary gland. Transgenic individuals are viable, fertile and transmit the rhEPO gene to the offspring. Northern blot data indicated that the expression of the transgene in the mammary gland is controlled by whey acidic protien (WAP) regulatory sequences during the period of lactation. While the hybridization with total RNA revealed the expression only in the lactating mammary gland, the highly sensitive combinatory approach using RT-PCR/hybridization technique detected a minor ectopic expression. The level of rhEPO secretion in the founder female, measured in the period of lactation, varied in the range of 60-178 and 60-162 mIU/ml in the milk and blood plasma, respectively. Biological activity of the milk rhEPO was confirmed by a standard [3H]-thymidine incorporation test. Thus, we describe the model of a rhEPO-transgenic rabbit, valuable for studies of rhEPO glycosylation and function, which can be useful for the development of transgenic approaches designed for the preparation of recombinant proteins by alternative biopharmaceutical production.

  4. Silencing erythropoietin receptor on glioma cells reinforces efficacy of temozolomide and X-rays through senescence and mitotic catastrophe.

    PubMed

    Pérès, Elodie A; Gérault, Aurélie N; Valable, Samuel; Roussel, Simon; Toutain, Jérôme; Divoux, Didier; Guillamo, Jean-Sébastien; Sanson, Marc; Bernaudin, Myriam; Petit, Edwige

    2015-02-10

    Hypoxia-inducible genes may contribute to therapy resistance in glioblastoma (GBM), the most aggressive and hypoxic brain tumours. It has been recently reported that erythropoietin (EPO) and its receptor (EPOR) are involved in glioma growth. We now investigated whether EPOR signalling may modulate the efficacy of the GBM current treatment based on chemotherapy (temozolomide, TMZ) and radiotherapy (X-rays). Using RNA interference, we showed on glioma cell lines (U87 and U251) that EPOR silencing induces a G2/M cell cycle arrest, consistent with the slowdown of glioma growth induced by EPOR knock-down. In vivo, we also reported that EPOR silencing combined with TMZ treatment is more efficient to delay tumour recurrence and to prolong animal survival compared to TMZ alone. In vitro, we showed that EPOR silencing not only increases the sensitivity of glioma cells to TMZ as well as X-rays but also counteracts the hypoxia-induced chemo- and radioresistance. Silencing EPOR on glioma cells exposed to conventional treatments enhances senescence and induces a robust genomic instability that leads to caspase-dependent mitotic death by increasing the number of polyploid cells and cyclin B1 expression. Overall these data suggest that EPOR could be an attractive target to overcome therapeutic resistance toward ionising radiation or temozolomide.

  5. The effect of hemin-induced oxidative stress on erythropoietin production in HepG2 cells.

    PubMed

    Nishimura, Kazuhiko; Tokida, Masahiro; Katsuyama, Hideaki; Nakagawa, Hiroshi; Matsuo, Saburo

    2014-11-01

    Erythropoietin (EPO) and iron are both indispensable hematopoietic factors and are often studied in humans and rodents. Iron activates prolyl hydroxylases (PHDs) and promotes the degradation of the α-subunit of hypoxia inducible factor (HIF), which regulates EPO production. Iron also causes oxidative stress. Oxidative stress leads to alterations in the levels of multiple factors that regulate HIF and EPO production. It is thought that iron influences EPO production by altering two pathways, namely PHDs activity and oxidative stress. We studied the differential effect of varying concentrations of hemin, an iron-containing porphyrin, on EPO production in HepG2 cells. Hemin at 100 µM reduced EPO mRNA expression. The hemin-induced reduction of EPO mRNA levels was attenuated at concentrations greater than 200 µM and EPO production increased in the presence of 500 µM hemin. In comparison, protoporphyrin IX, iron-free hemin did not influence EPO mRNA expression. Additionally, malondialdehyde (MDA) concentrations and superoxide dismutase (SOD) activity significantly increased with 300 µM hemin. Importantly, the antioxidant tempol inhibited the hemin-induced (500 µM) increase in EPO mRNA levels. In conclusion, these results suggest that restraint of EPO production by hemin was offset by the promotion of EPO production by hemin-induced oxidative stress at hemin concentrations greater than 300 µM.

  6. Inactivation of recombinant plasmid DNA from a human erythropoietin-producing mouse cell line grown on a large scale.

    PubMed

    Fibi, M R; Bröker, M; Schulz, R; Johannsen, R; Zettlmeissl, G

    1991-08-01

    Experiments were carried out to assess the survival of recombinant plasmid DNA during large-scale production of recombinant human erythropoietin (rhuEPO) in a fermentation pilot plant. The analyses revealed DNA-degrading activities in the fermentation broth and in the waste-water, leading to rapid destruction of plasmid DNA added to medium or waste-water. The capability of the plasmid-DNA-spiked samples to transform competent bacteria was drastically reduced. The DNA-degrading activity in the waste-waters could be blocked by addition of EDTA or by boiling, indicating the presence of DNA-degrading enzymes (DNases). No plasmid-specific DNA sequences were detected in waste-water samples by in-vitro amplification with Taq-polymerase. Genomic DNA preparations of cell debris collected from waste-water samples only contained degraded plasmid DNA. Furthermore, it was shown that intact plasmid DNA could be degraded to fragments of less than 1000 bp by incubation at 121 degrees C for 20 min, leading to a decrease in the plasmid-specific transforming capacity by a factor of 10(3) per minute. Thus, DNA from the rhuEPO production pilot plant was efficiently inactivated at three different levels: (i) in the fermentation medium (DNase), (ii) in the waste-water container (DNase), and (iii) by heat inactivation for 20 min at 120 degrees C. These results indicate that the probability of delivery of recombinant DNA into the environment is extremely low in such biotechnological production processes.

  7. Erythropoietin withdrawal alters interactions between young red blood cells, splenic endothelial cells, and macrophages: an in vitro model of neocytolysis

    NASA Technical Reports Server (NTRS)

    Trial, J.; Rice, L.; Alfrey, C. P.

    2001-01-01

    BACKGROUND: We have described the rapid destruction of young red blood cells (neocytolysis) in astronauts adapting to microgravity, in polycythemic high altitude dwellers who descend to sea level, and in patients with kidney disorders. This destruction results from a decrease in erythropoietin (EPO) production. We hypothesized that such EPO withdrawal could trigger physiological changes in cells other than red cell precursors and possibly lead to the uptake and destruction of young red cells by altering endothelial cell-macrophage interactions, most likely occurring in the spleen. METHODS: We identified EPO receptors on human splenic endothelial cells (HSEC) and investigated the responses of these cells to EPO withdrawal. RESULTS: A monolayer of HSEC, unlike human endothelial cells from aorta, glomerulus, or umbilical vein, demonstrated an increase in permeability upon EPO withdrawal that was accompanied by unique morphological changes. When HSEC were cultured with monocyte-derived macrophages (but not when either cell type was cultured alone), EPO withdrawal induced an increased ingestion of young red cells by macrophages when compared with the constant presence or absence of EPO. CONCLUSIONS: HSEC may represent a unique cell type that is able to respond to EPO withdrawal by increasing permeability and interacting with phagocytic macrophages, which leads to neocytolysis.

  8. Human erythropoietin induces a pro-angiogenic phenotype in cultured endothelial cells and stimulates neovascularization in vivo.

    PubMed

    Ribatti, D; Presta, M; Vacca, A; Ria, R; Giuliani, R; Dell'Era, P; Nico, B; Roncali, L; Dammacco, F

    1999-04-15

    Hematopoietic and endothelial cell lineages share common progenitors. Accordingly, cytokines formerly thought to be specific for the hematopoietic system have been shown to affect several functions in endothelial cells, including angiogenesis. In this study, we investigated the angiogenic potential of erythropoietin (Epo), the main hormone regulating proliferation, differentiation, and survival of erythroid cells. Epo receptors (EpoRs) have been identified in the human EA.hy926 endothelial cell line by Western blot analysis. Also, recombinant human Epo (rHuEpo) stimulates Janus Kinase-2 (JAK-2) phosphorylation, cell proliferation, and matrix metalloproteinase-2 (MMP-2) production in EA.hy926 cells and significantly enhances their differentiation into vascular structures when seeded on Matrigel. In vivo, rHuEpo induces a potent angiogenic response in the chick embryo chorioallantoic membrane (CAM). Accordingly, endothelial cells of the CAM vasculature express EpoRs, as shown by immunostaining with an anti-EpoR antibody. The angiogenic response of CAM blood vessels to rHuEpo was comparable to that elicited by the prototypic angiogenic cytokine basic fibroblast growth factor (FGF2), it occurred in the absence of a significant mononuclear cell infiltrate, and it was not mimicked by endothelin-1 (ET-1) treatment. Taken together, these data demonstrate the ability of Epo to interact directly with endothelial cells and to elicit an angiogenic response in vitro and in vivo and thus act as a bona fide direct angiogenic factor.

  9. Role of erythropoietin in the angiogenic activity of bone marrow endothelial cells of MGUS and multiple myeloma patients

    PubMed Central

    Ferrucci, Arianna; Ria, Roberto; Ruggieri, Simona; Racanelli, Vito; Rao, Luigia; Annese, Tiziana; Nico, Beatrice; Vacca, Angelo; Ribatti, Domenico

    2016-01-01

    Increasing evidences suggest several biological roles for erythropoietin and its receptor (Epo and EpoR), unrelated to erythropoiesis, including angiogenesis. Here, we detected the expression of EpoR in bone marrow-derived endothelial cells from monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM) patients (MGECs and MMECs, respectively) and assessed whether Epo plays a role in MGECs- and MMECs-mediated angiogenesis. We show that EpoR is expressed by both MGECs and MMECs even though at a higher level in the first ones. Both EC types respond to rHuEpo in terms of cell proliferation, whereas other responses, including activation of JAK2/STAT5 and PI3K/Akt pathways, cell migration and capillarogenesis are enhanced by Epo in MGECs, but not in MMECs. In addition, the conditioned media of both Epo-treated cells induce a strong angiogenic response in vivo in the chorioallantoic membrane assay, comparable to that of vascular endothelial growth factor (VEGF). Overall, these data highlight the effect of Epo on MGECs- and MMECs-mediated angiogenesis: MGECs are more responsive to Epo treatment than MMECs, probably because over-angiogenic phenotype of MMECs is already activated by their autocrine/paracrine loops occurring in the “angiogenic switch” from MGUS. PMID:26919105

  10. Erythropoietin improves neuronal proliferation in dentate gyrus of hippocampal formation in an animal model of Alzheimer's disease

    PubMed Central

    Arabpoor, Zohreh; Hamidi, Gholamali; Rashidi, Bahman; Shabrang, Moloud; Alaei, Hojjatallah; Sharifi, Mohammad Reza; Salami, Mahmoud; Dolatabadi, Hamid Reza Dehghani; Reisi, Parham

    2012-01-01

    Background: Alzheimer's disease (AD) is a prevalent disorder with severe learning and memory defects. Because it has been demonstrated that erythropoietin (EPO) has positive effects on the central nervous system, the aim of this study was to evaluate the effect of EPO on neuronal proliferation in dentate gyrus of hippocampal formation in a well-defined model for AD. Materials and Methods: A rat model of sporadic dementia of Alzheimer's type was established by a bilateral intracerebroventricular injection of streptozotocin (ICV-STZ). Impairment of learning and memory was confirmed 2 weeks after ICV-STZ injection by passive avoidance learning test and then rats were divided into fourgroups:Control, control-EPO, Alzheimer and Alzheimer-EPO. EPO was injected intraperitoneally every other day with a dose of 5000 IU/kg and, finally, the rats were anesthetized and decapitated for immunohistochemical study and neurogenesis investigation (by Ki67 method) in dentate gyrus of hippocampal formation. Results: The results driven from the histological study showed that EPO significantly increases neuronal proliferation in dentate gyrus of hippocampus in the Alzheimer-EPO group compared with the control, control-EPO and Alzheimer groups; however, there were no differences between the other groups. Conclusion: Our results show that even though EPO in intact animals doesnot change neurogenesis in dentate gyrus, it can nonetheless significantly increase neurogenesis if there is an underlying disorder like neurodegenerative diseases. PMID:23326781

  11. Erythropoietin Levels Increase during Cerebral Malaria and Correlate with Heme, Interleukin-10 and Tumor Necrosis Factor-Alpha in India

    PubMed Central

    Dalko, Esther; Tchitchek, Nicolas; Pays, Laurent; Herbert, Fabien; Cazenave, Pierre-André; Ravindran, Balachandran; Sharma, Shobhona; Nataf, Serge; Das, Bidyut; Pied, Sylviane

    2016-01-01

    Cerebral malaria (CM) caused by Plasmodium falciparum parasites often leads to the death of infected patients or to persisting neurological sequelae despite anti-parasitic treatments. Erythropoietin (EPO) was recently suggested as a potential adjunctive treatment for CM. However diverging results were obtained in patients from Sub-Saharan countries infected with P. falciparum. In this study, we measured EPO levels in the plasma of well-defined groups of P. falciparum-infected patients, from the state of Odisha in India, with mild malaria (MM), CM, or severe non-CM (NCM). EPO levels were then correlated with biological parameters, including parasite biomass, heme, tumor necrosis factor (TNF)-α, interleukin (IL)-10, interferon gamma-induced protein (IP)-10, and monocyte chemoattractant protein (MCP)-1 plasma concentrations by Spearman’s rank and multiple correlation analyses. We found a significant increase in EPO levels with malaria severity degree, and more specifically during fatal CM. In addition, EPO levels were also found correlated positively with heme, TNF-α, IL-10, IP-10 and MCP-1 during CM. We also found a significant multivariate correlation between EPO, TNF-α, IL-10, IP-10 MCP-1 and heme, suggesting an association of EPO with a network of immune factors in CM patients. The contradictory levels of circulating EPO reported in CM patients in India when compared to Africa highlights the need for the optimization of adjunctive treatments according to the targeted population. PMID:27441662

  12. Erythropoietin and a nonerythropoietic peptide analog promote aortic endothelial cell repair under hypoxic conditions: role of nitric oxide

    PubMed Central

    Heikal, Lamia; Ghezzi, Pietro; Mengozzi, Manuela; Stelmaszczuk, Blanka; Feelisch, Martin; Ferns, Gordon AA

    2016-01-01

    The cytoprotective effects of erythropoietin (EPO) and an EPO-related nonerythropoietic analog, pyroglutamate helix B surface peptide (pHBSP), were investigated in an in vitro model of bovine aortic endothelial cell injury under normoxic (21% O2) and hypoxic (1% O2) conditions. The potential molecular mechanisms of these effects were also explored. Using a model of endothelial injury (the scratch assay), we found that, under hypoxic conditions, EPO and pHBSP enhanced scratch closure by promoting cell migration and proliferation, but did not show any effect under normoxic conditions. Furthermore, EPO protected bovine aortic endothelial cells from staurosporine-induced apoptosis under hypoxic conditions. The priming effect of hypoxia was associated with stabilization of hypoxia inducible factor-1α, EPO receptor upregulation, and decreased Ser-1177 phosphorylation of endothelial nitric oxide synthase (NOS); the effect of hypoxia on the latter was rescued by EPO. Hypoxia was associated with a reduction in nitric oxide (NO) production as assessed by its oxidation products, nitrite and nitrate, consistent with the oxygen requirement for endogenous production of NO by endothelial NOS. However, while EPO did not affect NO formation in normoxia, it markedly increased NO production, in a manner sensitive to NOS inhibition, under hypoxic conditions. These data are consistent with the notion that the tissue-protective actions of EPO-related cytokines in pathophysiological settings associated with poor oxygenation are mediated by NO. These findings may be particularly relevant to atherogenesis and postangioplasty restenosis. PMID:27800514

  13. The effect of hemin-induced oxidative stress on erythropoietin production in HepG2 cells.

    PubMed

    Nishimura, Kazuhiko; Tokida, Masahiro; Katsuyama, Hideaki; Nakagawa, Hiroshi; Matsuo, Saburo

    2014-11-01

    Erythropoietin (EPO) and iron are both indispensable hematopoietic factors and are often studied in humans and rodents. Iron activates prolyl hydroxylases (PHDs) and promotes the degradation of the α-subunit of hypoxia inducible factor (HIF), which regulates EPO production. Iron also causes oxidative stress. Oxidative stress leads to alterations in the levels of multiple factors that regulate HIF and EPO production. It is thought that iron influences EPO production by altering two pathways, namely PHDs activity and oxidative stress. We studied the differential effect of varying concentrations of hemin, an iron-containing porphyrin, on EPO production in HepG2 cells. Hemin at 100 µM reduced EPO mRNA expression. The hemin-induced reduction of EPO mRNA levels was attenuated at concentrations greater than 200 µM and EPO production increased in the presence of 500 µM hemin. In comparison, protoporphyrin IX, iron-free hemin did not influence EPO mRNA expression. Additionally, malondialdehyde (MDA) concentrations and superoxide dismutase (SOD) activity significantly increased with 300 µM hemin. Importantly, the antioxidant tempol inhibited the hemin-induced (500 µM) increase in EPO mRNA levels. In conclusion, these results suggest that restraint of EPO production by hemin was offset by the promotion of EPO production by hemin-induced oxidative stress at hemin concentrations greater than 300 µM. PMID:24962609

  14. Translation of the human erythropoietin transcript is regulated by an upstream open reading frame in response to hypoxia.

    PubMed

    Barbosa, Cristina; Romão, Luísa

    2014-05-01

    Erythropoietin (EPO) is a key mediator hormone for hypoxic induction of erythropoiesis that also plays important nonhematopoietic functions. It has been shown that EPO gene expression regulation occurs at different levels, including transcription and mRNA stabilization. In this report, we show that expression of EPO is also regulated at the translational level by an upstream open reading frame (uORF) of 14 codons. As judged by comparisons of protein and mRNA levels, the uORF acts as a cis-acting element that represses translation of the main EPO ORF in unstressed HEK293, HepG2, and HeLa cells. However, in response to hypoxia, this repression is significantly released, specifically in HeLa cells, through a mechanism that involves processive scanning of ribosomes from the 5' end of the EPO transcript and enhanced ribosome bypass of the uORF. In addition, we demonstrate that in HeLa cells, hypoxia induces the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α) concomitantly with a significant increase of EPO protein synthesis. These findings provide a framework for understanding that production of high levels of EPO induced by hypoxia also involves regulation at the translational level. PMID:24647661

  15. TNF-alpha-induced apoptosis is prevented by erythropoietin treatment on SH-SY5Y cells

    SciTech Connect

    Pregi, Nicolas Wenker, Shirley; Vittori, Daniela; Leiros, Claudia Perez; Nesse, Alcira

    2009-02-01

    The growth factor erythropoietin (Epo) has shown neuronal protective action in addition to its well known proerythroid activity. Furthermore, Epo has dealt with cellular inflammation by inhibiting the expression of several proinflammatory cytokines, such as IL-1 and TNF-{alpha}. The action of TNF can have both apoptotic and antiapoptotic consequences due to altered balance between different cell signalling pathways. This work has focused on the apoptotic effects of this cytokine and the potential protective action of Epo. The model we used was neuroblastoma SH-SY5Y cells cultured in the presence of 25 ng/ml TNF-{alpha} or pretreated with 25 U/ml Epo for 12 h before the addition of TNF-{alpha}. Apoptosis was evaluated by differential cell count after Hoechst staining, analysis of DNA ladder pattern, and measurement of caspase activity. Despite its ability to induce NF-{kappa}B nuclear translocation, TNF-{alpha} induced cell death, which was found to be associated to upregulation of TNF Receptor 1 expression. On the other hand, cells activated by Epo became resistant to cell death. Prevention of death receptor upregulation and caspase activation may explain this antiapoptotic effect of Epo, which may be also favoured by the induction of a higher expression of protective factors, such as Bcl-2 and NF-{kappa}B, through mechanisms involving Jak/STAT and PI3K signalling pathways.

  16. Veterans Administration Databases

    Cancer.gov

    The Veterans Administration Information Resource Center provides database and informatics experts, customer service, expert advice, information products, and web technology to VA researchers and others.

  17. Investigation the efficacy of intra-articular prolotherapy with erythropoietin and dextrose and intra-articular pulsed radiofrequency on pain level reduction and range of motion improvement in primary osteoarthritis of knee

    PubMed Central

    Rahimzadeh, Poupak; Imani, Farnad; Faiz, Seyed Hamid Reza; Entezary, Saeed Reza; Nasiri, Ali Akbar; Ziaeefard, Mohsen

    2014-01-01

    Background: Osteoarthritis is one of the most common diseases and the knee is the most commonly affected joint. Intra-articular prolotherapy is being utilized in acute and chronic pain management setting. This study was designed to compare the efficacy of three methods of intra-articular knee joint therapies with erythropoietin, dextrose, and pulsed radiofrequency. Materials and Methods: After approval by the Ethics Committee and explaining the therapeutic method to volunteers, 70 patients who were suffering from primary knee osteoarthrosis went through one of the treatment methods (erythropoietin, dextrose, and pulsed radiofrequency). The study was double-blind randomized clinical trial performed from December 2012 to July 2013. Patients’ pain level was assessed through the visual analog pain scale (VAS), and range of motion (ROM) was measured by goniometric method. Furthermore, patients’ satisfaction was assessed before and after different treatment methods in weeks 2, 4, and 12. For analysis, Chi-square, one-way ANOVA, and repeated measured ANOVA were utilized. Results: The demographic results among the three groups did not indicate any statistical difference. The mean VAS in erythropoietin group in the 2nd, 4th, and 12th weeks was 3.15 ± 1.08, 3.15 ± 1.08, and 3.5 ± 1.23, respectively (P ≤ 0.005). Knee joint ROM in the erythropoietin group in the 2nd, 4th, and 12th weeks was 124 ± 1.50, 124 ± 1.4, and 123 ± 1.53 respectively (P ≤ 0.005). Satisfaction score in the 12th week in erythropoietin group was extremely satisfied 15%, satisfied 55%, and moderately satisfied 30%, (P = 0.005). No specific side-effects were observed. Conclusion: Intra-articular prolotherapy with erythropoietin was more effective in terms of pain level reduction and ROM improvement compared with dextrose and pulsed radiofrequency. PMID:25422652

  18. Pharmacokinetics and selected pharmacodynamics of cobalt following a single intravenous administration to horses.

    PubMed

    Knych, H K; Arthur, R M; Mitchell, M M; Holser, I; Poppenga, R; Smith, L L; Helm, M N; Sams, R A; Gaskill, C L

    2015-07-01

    Cobalt has been used by human athletes due to its purported performance-enhancing effects. It has been suggested that cobalt administration results in enhanced erythropoiesis, secondary to increased circulating erythropoietin (EPO) concentrations leading to improvements in athletic performance. Anecdotal reports of illicit administration of cobalt to horses for its suspected performance enhancing effects have led us to investigate the pharmacokinetics and pharmacodynamic effects of this compound when administered in horses, so as to better regulate its use. In the current study, 18 horses were administered a single intravenous dose of cobalt chloride or cobalt gluconate and serum and urine samples collected for up to 10 days post administration. Cobalt concentrations were measured using inductively coupled plasma mass spectrometry (ICP-MS) and pharmacokinetic parameters determined. Additional blood samples were collected for measurement of equine EPO concentrations as well as to assess any effects on red blood cell parameters. Horses were observed for adverse effects and heart rate monitored for the first 4 h post administration. Cobalt was characterized by a large volume of distribution (0.939 L/kg) and a prolonged gamma half-life (156.4 h). Cobalt serum concentrations were still above baseline values at 10 days post administration. A single administration of cobalt had no effect on EPO concentrations, red blood cell parameters or heart rate in any of the horses studied and no adverse effects were noted. Based on the prolonged gamma half-life and prolonged residence time, regulators should be able to detect administration of a single dose of cobalt to horses.

  19. The Administrative Power Grab

    ERIC Educational Resources Information Center

    Sorenson, Richard D.

    2007-01-01

    Administrative power for some school teachers can be an aphrodisiac that can be applied negatively, especially when a leader has devastating instinct for the weaknesses of others. A leader's intellect and heart closes shop and ceases to function when drunk on power. In this article, the author describes how the use of administrative power can be…

  20. Innovation in Administrator Preparation.

    ERIC Educational Resources Information Center

    Coleman, Don

    An innovative administrator preparation program based on guidelines established by national boards and commissions is described in this paper. The California State University, Fresno, administrator education curriculum was reorganized by faculty and an advisory committee of 11 superintendents to meet the needs of local school districts. The…

  1. Migrant Education Administrative Guide.

    ERIC Educational Resources Information Center

    North Carolina State Dept. of Public Instruction, Raleigh. Div. of Compensatory Education.

    Relating specifically to the North Carolina migrant education program's administrative responsibilities, this guide is designed to aid administrators in program management, monitoring project activities, project evaluation, self-assessment, determining needs for training and staff development, site-visit preparation, policy development, and…

  2. Administration of Computer Resources.

    ERIC Educational Resources Information Center

    Franklin, Gene F.

    Computing at Stanford University has, until recently, been performed at one of five facilities. The Stanford hospital operates an IBM 370/135 mainly for administrative use. The university business office has an IBM 370/145 for its administrative needs and support of the medical clinic. Under the supervision of the Stanford Computation Center are…

  3. Issues in Educational Administration.

    ERIC Educational Resources Information Center

    Ediger, Marlow

    School administrators need to study and analyze the pros and cons of issues before making decisions. Ultimately, decisions need to be made by administrators as to which philosophies of education to implement in resolving conflicting points of view. More research studies would lead to an increased number of syntheses of the pros and cons of certain…

  4. Justifying Educational Administration.

    ERIC Educational Resources Information Center

    Evers, Colin; Lakomski, Gabriele

    1993-01-01

    The traditional conceptions of science dominating educational administration are mistaken. Unacceptable epistemologies, like those implicit in logical positivism, justify knowledge solely in terms of empirical adequacy. An improved science of educational administration embraces a coherent global theory accounting for all the phenomena of human…

  5. Test Administration Models

    ERIC Educational Resources Information Center

    Becker, Kirk A.; Bergstrom, Betty A.

    2013-01-01

    The need for increased exam security, improved test formats, more flexible scheduling, better measurement, and more efficient administrative processes has caused testing agencies to consider converting the administration of their exams from paper-and-pencil to computer-based testing (CBT). Many decisions must be made in order to provide an optimal…

  6. Champions of Children. Administrators . . .

    ERIC Educational Resources Information Center

    Chaffee, John; Olds, H. Robert

    Today, in an era of taxpayer revolts, lack of clarity in values, and changing family structure, children need advocates in the political arena as well as in the schools. This pamphlet suggests that administrators are in an excellent position to defend the rights of children on all fronts. It focuses on what administrators have done and specific…

  7. The School Personnel Administrator.

    ERIC Educational Resources Information Center

    Knox, Rodney F.

    This paper provides an overview of the development of the school-personnel administrator role. It first describes the influence of the science-management and human-relations movements and the behavioral sciences on personnel administration and human resource management. It next discusses the role of the personnel-performance-appraisal system and…

  8. Tenure for Administrators?

    ERIC Educational Resources Information Center

    Holyer, Robert

    2004-01-01

    The practice of granting tenure to academic administrators, especially presidents and academic deans, seems fairly prevalent. However, it is important to consider the possible advantages and disadvantages carefully before making such an offer. Boards interested in attracting talented administrators empowered to do what is in the best interest of…

  9. 47 CFR 54.715 - Administrative expenses of the Administrator.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 47 Telecommunication 3 2011-10-01 2011-10-01 false Administrative expenses of the Administrator. 54.715 Section 54.715 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) COMMON CARRIER SERVICES (CONTINUED) UNIVERSAL SERVICE Administration § 54.715 Administrative expenses of the Administrator. (a) The annual administrative expenses...

  10. Analytics of nonpeptidic erythropoietin mimetic agents in sports drug testing employing high-resolution/high-accuracy liquid chromatography-mass spectrometry.

    PubMed

    Vogel, Matthias; Dib, Josef; Tretzel, Laura; Piper, Thomas; Thomas, Andreas; Schänzer, Wilhelm; Thevis, Mario

    2016-09-01

    Since its release as anti-anemic drug, recombinant erythropoietin (rEPO) gradually entered the illicit way to sports competitions as endurance-enhancing drug. Novel modifications biopharmaceutically introduced into the rEPO molecule in the form of carbohydrate or polyethylene glycol moieties made robust and sensitive test methods vital to doping controls in order to provide the necessary tools enabling the conviction of dishonest athletes. Modern protein analysis by means of gel electrophoretic separation and western blotting represents the status quo in rEPO anti-doping analysis. However, new therapeutically promising erythropoietin receptor activating compounds have been developed that exhibit cytokine hormone-mimicking properties but lack any protein structure. Progression to evade parenteral application and substitute for rEPO by low molecular mass and orally available compounds is still one of the major objectives in pharmaceutical research. In this approach, four promising in-house synthesized nonpeptidic erythropoietin mimetic agents, namely compound 129, compound 163, A1B10C1, and A5B10C4 were thoroughly evaluated by employing high-resolution/high-accuracy liquid chromatography tandem mass spectrometry experiments. Characteristic product ions were determined supporting the identification of these drugs and putative metabolites as well as related compounds in future doping controls. Test methods employing direct urine injection and receptor affinity purification strategies were assessed, which demonstrated that EPO receptor purification is of limited utility for nonpeptidic EPOR agonists while direct urine injection allowed for comprehensive method characterization. Thereby, achieved limits of detection were 1 ng/mL for compounds 129/163 and 5 ng/mL for A1B10C1/A5B10C4. PMID:27438721

  11. Hormone dependency of chromosome aberrations induced by 7,12-dimethylbenz(a)anthracene in rat bone marrow cells: site-specific increase by erythropoietin

    SciTech Connect

    Ueda, N.; Suglyama, T.; Chattopadhyay, S.C.; Goto-Mimura, K.; Maeda, S.

    1981-08-01

    The frequency of chromosome aberrations (CA) 6 hours after iv injection of 50 mg 7,12-dimethylbenz(a)anthracene (DMBA0/kg was studied in bone marrow cells of the noninbred Long-Evans rat under various hematopoietic conditions. The percentage of metaphase cells with CA was enhanced by anemia and suppressed by polycythemia. The low incidence of CA in polycythemic rats was reversed by 6 U of sheep erythropoietin (EP) injected at the time of DMBA treatment. The interchromosomal and intrachromosomal distribution of CA indicated that hematopoietic stimuli, more specifically EP, greatly enhanced DMBA-induced CA in specific chromosomal regions.

  12. The erythropoietin receptor is not required for the development, function, and aging of rods and cells in the retinal periphery

    PubMed Central

    Caprara, Christian; Britschgi, Corinne; Samardzija, Marijana

    2014-01-01

    Purpose Erythropoietin (EPO) was originally described for its antiapoptotic effects on erythroid progenitor cells in bone marrow. In recent years, however, EPO has also been shown to be cytoprotective in several tissues, including the retina. There, exogenous application of EPO was reported to exert neuro- and vasoprotection in several models of retinal injury. EPO and the erythropoietin receptor (EPOR) are expressed in the retina, but the role of endogenous EPO-EPOR signaling in this tissue remains elusive. Here, we investigated the consequences for cell physiology and survival when EpoR is ablated in rod photoreceptors or in the peripheral retina. Methods Two mouse lines were generated harboring a cyclization recombinase (CRE)-mediated knockdown of EpoR in rod photoreceptors (EpoRflox/flox;Opn-Cre) or in a heterogeneous cell population of the retinal periphery (EpoRflox/flox;α-Cre). The function of the retina was measured with electroretinography. Retinal morphology was analyzed in tissue sections. The vasculature of the retina was investigated on flatmount preparations, cryosections, and fluorescein angiography. Retinal nuclear layers were isolated by laser capture microdissection to test for EpoR expression. Gene expression analysis was performed with semiquantitative real-time PCR. To test if the absence of EPOR potentially increases retinal susceptibility to hypoxic stress, the knockdown mice were exposed to hypoxia. Results Newborn mice had lower retinal expression levels of EpoR and soluble EpoR (sEpoR) than the adult wild-type mice. In the adult mice, the EpoR transcripts were elevated in the inner retinal layers, while expression in the photoreceptors was low. CRE-mediated deletion in the EpoRflox/flox;Opn-Cre mice led to a decrease in EpoR mRNA expression in the outer nuclear layer. A significant decrease in EpoR expression was measured in the retina of the EpoRflox/flox;α-Cre mice, accompanied by a strong and significant decrease in sEpoR expression

  13. Limiting hepatic Bmp-Smad signaling by matriptase-2 is required for erythropoietin-mediated hepcidin suppression in mice

    PubMed Central

    Nai, Antonella; Rubio, Aude; Campanella, Alessandro; Gourbeyre, Ophélie; Artuso, Irene; Bordini, Jessica; Gineste, Aurélie; Latour, Chloé; Besson-Fournier, Céline; Lin, Herbert Y.; Coppin, Hélène; Roth, Marie-Paule; Camaschella, Clara; Silvestri, Laura

    2016-01-01

    Hepcidin, the main regulator of iron homeostasis, is repressed when erythropoiesis is acutely stimulated by erythropoietin (EPO) to favor iron supply to maturing erythroblasts. Erythroferrone (ERFE) has been identified as the erythroid regulator that inhibits hepcidin in stress erythropoiesis. A powerful hepcidin inhibitor is the serine protease matriptase-2, encoded by TMPRSS6, whose mutations cause iron refractory iron deficiency anemia. Because this condition has inappropriately elevated hepcidin in the presence of high EPO levels, a role is suggested for matriptase-2 in EPO-mediated hepcidin repression. To investigate the relationship between EPO/ERFE and matriptase-2, we show that EPO injection induces Erfe messenger RNA expression but does not suppress hepcidin in Tmprss6 knockout (KO) mice. Similarly, wild-type (WT) animals, in which the bone morphogenetic protein–mothers against decapentaplegic homolog (Bmp-Smad) pathway is upregulated by iron treatment, fail to suppress hepcidin in response to EPO. To further investigate whether the high level of Bmp-Smad signaling of Tmprss6 KO mice counteracts hepcidin suppression by EPO, we generated double KO Bmp6-Tmprss6 KO mice. Despite having Bmp-Smad signaling and hepcidin levels that are similar to WT mice under basal conditions, double KO mice do not suppress hepcidin in response to EPO. However, pharmacologic downstream inhibition of the Bmp-Smad pathway by dorsomorphin, which targets the BMP receptors, improves the hepcidin responsiveness to EPO in Tmprss6 KO mice. We concluded that the function of matriptase-2 is dominant over that of ERFE and is essential in facilitating hepcidin suppression by attenuating the BMP-SMAD signaling. PMID:26755707

  14. Serum Proteomic Changes after Randomized Prolonged Erythropoietin Treatment and/or Endurance Training: Detection of Novel Biomarkers

    PubMed Central

    Christensen, Britt; Ludvigsen, Maja; Nellemann, Birgitte; Kopchick, John J.; Honoré, Bent; Jørgensen, Jens Otto L.

    2015-01-01

    Introduction Despite implementation of the biological passport to detect erythropoietin abuse, a need for additional biomarkers remains. We used a proteomic approach to identify novel serum biomarkers of prolonged erythropoiesis-stimulating agent (ESA) exposure (Darbepoietin-α) and/or aerobic training. Trial Design Thirty-six healthy young males were randomly assigned to the following groups: Sedentary-placebo (n = 9), Sedentary-ESA (n = 9), Training-placebo (n = 10), or Training-ESA (n = 8). They were treated with placebo/Darbepoietin-α subcutaneously once/week for 10 weeks followed by a 3-week washout period. Training consisted of supervised biking 3/week for 13 weeks at the highest possible intensity. Serum was collected at baseline, week 3 (high dose Darbepoietin-α), week 10 (reduced dose Darbepoietin-α), and after a 3-week washout period. Methods Serum proteins were separated according to charge and molecular mass (2D-gel electrophoresis). The identity of proteins from spots exhibiting altered intensity was determined by mass spectrometry. Results Six protein spots changed in response to Darbepoietin-α treatment. Comparing all 4 experimental groups, two protein spots (serotransferrin and haptoglobin/haptoglobin related protein) showed a significant response to Darbepoietin-α treatment. The haptoglobin/haptoglobin related protein spot showed a significantly lower intensity in all subjects in the training-ESA group during the treatment period and increased during the washout period. Conclusion An isoform of haptoglobin/haptoglobin related protein could be a new anti-doping marker and merits further research. Trial Registration ClinicalTrials.gov NCT01320449 PMID:25679398

  15. Mutations in the WSAWSE and cytosolic domains of the erythropoietin receptor affect signal transduction and ligand binding and internalization.

    PubMed Central

    Quelle, D E; Quelle, F W; Wojchowski, D M

    1992-01-01

    The terminal development of erythroid progenitor cells is promoted in part through the interaction of erythropoietin (EPO) with its cell surface receptor. This receptor and a growing family of related cytokine receptors share homologous extracellular features, including a well-conserved WSXWS motif. To explore the functional significance of this motif in the murine EPO receptor, five WSAWSE mutants were prepared and their signal-transducing, ligand binding, and endocytotic properties were compared. EPO receptors mutated at tryptophan residues (W-232, W-235----G; W-235----G; W-235----F) failed to mediate EPO-induced growth or pp100 phosphorylation, while S-236----T and E-237----K mutants exhibited partial to full activity (50 to 100% of wild-type growth and induced phosphorylation). Ligand affinity was reduced for mutant receptors (two- to fivefold), yet expression at the cell surface for all receptors was nearly equivalent. Also, the ability of mutated receptors to internalize ligand was either markedly reduced or abolished (W-235----F), indicating a role for the WSAWSE region in hormone internalization. Interestingly, receptor forms lacking 97% of the cytosolic domain (no signal-transducing capacity; binding affinity reduced two- to threefold) internalized EPO efficiently. This and all WSAWSE receptor forms studied also mediated specific cross-linking of 125I-EPO to three accessory membrane proteins (M(r)s, 120,000, 105,000, and 93,000). These findings suggest that the WSAWSE domain of the EPO receptor is important for EPO-induced signal transduction and ligand internalization. In contrast, although the cytosolic domain is required for growth signaling, it appears nonessential for efficient endocytosis. Images PMID:1406645

  16. Sialic acid-specific affinity chromatography for the separation of erythropoietin glycoforms using serotonin as a ligand.

    PubMed

    Meininger, M; Stepath, M; Hennig, R; Cajic, S; Rapp, E; Rotering, H; Wolff, M W; Reichl, U

    2016-02-15

    Recombinant human erythropoietin (rhEPO) is an important CHO cell-derived glycoprotein and the degree of sialylation of this hormone is crucial for its in vivo bioactivity. In order to improve the purification process serotonin as a potential affinity ligand was tested for preparative chromatographic separation of rhEPO glycoforms into fractions of different degrees of sialylation. Therefore, two chromatographic matrices were prepared by immobilizing serotonin on CNBr- and NHS-Sepharose™. First it was shown both matrices bind rhEPO only in its sialylated form. Results indicate that binding is pH independent between pH 3.5 to 8 suggesting it is not only based on electrostatic interactions. Second, after optimal binding conditions were identified, semi-purified rhEPO was loaded onto both matrices and eluted using a stepwise elution gradient of sodium chloride. For comparison same affinity purification experiments were performed using wheat germ agglutinin-coupled agarose, a lectin known for its affinity towards sialylated glycoproteins. To monitor changes in N-glycan fingerprint, eluate fractions were analyzed by multiplexed capillary gel electrophoresis coupled to laser-induced fluorescence (xCGE-LIF). For the serotonin matrices an increasing degree of sialylation was observed from the first to the third elution fraction while purity of rhEPO could be increased at the same time. The late elution fractions of serotonin-coupled CNBr- and NHS-Sepharose™ also showed an overall sialylation degree exceeding that of the starting material. In contrast, for rhEPO bound to wheat germ agglutinin-coupled agarose, no distinct change in the degree of sialylation could be observed after elution. Overall, these encouraging results highlight the potential of serotonin as a chromatographic ligand for the improvement of pharmaceutical purification processes of rhEPO.

  17. Recombinant human erythropoietin and the quality of life of end-stage renal disease patients: a comparative analysis.

    PubMed

    Evans, R W

    1991-10-01

    The clinical and quality of life outcomes of hemodialysis patients improve remarkably following treatment with recombinant human erythropoietin (Epo). However, few studies have compared the quality of life of Epo patients with that of end-stage renal disease (ESRD) patients on various treatment modalities. Data obtained in three separate studies of ESRD patients were comparatively analyzed. Over 1,500 patients from 23 dialysis and transplant centers were studied. Both objective and subjective quality of life were examined. Objective quality of life indicators included employment status, functional ability, and health status. Subjective quality of life indicators included well-being, life satisfaction, psychological affect, and happiness. Quality of life varied significantly across treatment modality, with transplant recipients generally reporting the highest levels of objective and subjective quality of life. However, hemodialysis patients treated with Epo reported a statistically significant improvement between baseline and 10 months' follow-up on all quality of life indicators, except employment. Epo patients reported a level of overall life satisfaction that exceeded that of patients on all ESRD treatment modalities. Among transplant recipients, diabetics reported the poorest quality of life, while patients on conventional immunosuppressive therapy often had a quality of life that exceeded that of patients on cyclosporine therapy. Some of these findings may be explained by case-mix differences, as well as differing study designs. Quality of life remains a significant concern among ESRD patients and the physicians and medical professionals responsible for their care. Unfortunately, the rehabilitation potential of many patients, despite the availability of Epo, and the success of transplantation, remains unmet. PMID:1928082

  18. The effect of high altitude on platelet counts, thrombopoietin and erythropoietin levels in young Bolivian airmen visiting the Andes

    NASA Astrophysics Data System (ADS)

    Hudson, J. G.; Bowen, Angela L.; Navia, Pilar; Rios-Dalenz, Jaime; Pollard, Andrew J.; Williams, David; Heath, Donald

    Recognition of thrombosis as a complication of exposure to high altitude has stimulated interest in rheological changes resulting from hypobaric hypoxia. Previous studies of platelet counts at high altitude have yielded conflicting results and have not been studied in conjunction with potential mediating cytokines. We studied the effects of high-altitude exposure on platelet numbers, thrombopoietin (tpo) and erythropoietin (epo) levels in man. A group of 28 volunteers from the Bolivian Airforce stationed at Santa Cruz (600 m altitude) were studied 48 h and 1 week after their ascent to La Paz (3600 m). In addition 105 volunteers based at Santa Cruz for at least 1 year were compared with 175 age- and sex-matched residents at El Alto (4200 m). Platelet counts were measured immediately after sampling and serum samples assayed for tpo and epo. In the ascending group, mean platelet counts were 251×109, 367×109 and 398×109/l at 600 m and following 48 h and 1 week at 3600 m respectively. Mean tpo levels were 132.5, 76 and 92 pg/ml with epo values of 2.98, 11.6 and 7.9 mIU/ml respectively. In the resident populations mean platelet counts were 271×109/l in the low- and 471×109/l in the high-altitude groups. Mean tpo and epo levels measured 69.3 pg/ml and 4.5 mIU/ml respectively at 600 m and 58.5 pg/ml and 5.1 mIU/ml at 4200 m. In conclusion we have demonstrated a significant and sustained elevation in platelet numbers within 48 h of ascent to high altitude. Our findings do not support a role for tpo as a mediator of the increased platelet count. However, these data do not discount epo as a potential candidate.

  19. Erythropoietin Ameliorates Neonatal Hypoxia-Ischemia-Induced Neurobehavioral Deficits, Neuroinflammation, and Hippocampal Injury in the Juvenile Rat

    PubMed Central

    Lan, Kuo-Mao; Tien, Lu-Tai; Cai, Zhengwei; Lin, Shuying; Pang, Yi; Tanaka, Sachiko; Rhodes, Philip G.; Bhatt, Abhay J.; Savich, Renate D.; Fan, Lir-Wan

    2016-01-01

    The hematopoietic growth factor erythropoietin (EPO) has been shown to be neuroprotective against hypoxia-ischemia (HI) in Postnatal Day 7 (P7)–P10 or adult animal models. The current study was aimed to determine whether EPO also provides long-lasting neuroprotection against HI in P5 rats, which is relevant to immature human infants. Sprague-Dawley rats at P5 were subjected to right common carotid artery ligation followed by an exposure to 6% oxygen with balanced nitrogen for 1.5 h. Human recombinant EPO (rEPO, at a dose of 5 units/g) was administered intraperitoneally one hour before or immediately after insult, followed by additional injections at 24 and 48 h post-insult. The control rats were injected with normal saline following HI. Neurobehavioral tests were performed on P8 and P20, and brain injury was examined on P21. HI insult significantly impaired neurobehavioral performance including sensorimotor, locomotor activity and cognitive ability on the P8 and P20 rats. HI insult also resulted in brain inflammation (as indicated by microglia activation) and neuronal death (as indicated by Jade B positive staining) in the white matter, striatum, cortex, and hippocampal areas of the P21 rat. Both pre- and post-treatment with rEPO significantly improved neurobehavioral performance and protected against the HI-induced neuronal death, microglia activation (OX42+) as well as loss of mature oligodendrocytes (APC-CC1+) and hippocampal neurons (Nissl+). The long-lasting protective effects of rEPO in the neonatal rat HI model suggest that to exert neurotrophic activity in the brain might be an effective approach for therapeutic treatment of neonatal brain injury induced by hypoxia-ischemia. PMID:26927081

  20. Active conformation of the erythropoietin receptor: random and cysteine-scanning mutagenesis of the extracellular juxtamembrane and transmembrane domains.

    PubMed

    Lu, Xiaohui; Gross, Alec W; Lodish, Harvey F

    2006-03-17

    In the absence of erythropoietin (Epo) cell surface Epo receptors (EpoR) are dimeric; dimerization is mediated mainly by the transmembrane domain. Binding of Epo changes the orientation of the two receptor subunits. This conformational change is transmitted through the juxtamembrane and transmembrane domains, leading to activation of JAK2 kinase and induction of proliferation and survival signals. To define the active EpoR conformation(s) we screened libraries of EpoRs with random mutations in the transmembrane domain and identified several point mutations that activate the EpoR in the absence of ligand, including changes of either of the first two transmembrane domain residues (Leu(226) and Ile(227)) to cysteine. Following this discovery, we performed cysteine-scanning mutagenesis in the EpoR juxtamembrane and transmembrane domains. Many mutants formed disulfide-linked receptor dimers, but only EpoR dimers linked by cysteines at positions 223, 226, or 227 activated EpoR signal transduction pathways and supported proliferation of Ba/F3 cells in the absence of cytokines. These data suggest that activation of dimeric EpoR by Epo binding is achieved by reorienting the EpoR transmembrane and the connected cytosolic domains and that certain disulfide-bonded dimers represent the activated dimeric conformation of the EpoR, constitutively activating downstream signaling. Based on our data and the previously determined structure of Epo bound to a dimer of the EpoR extracellular domain, we present a model of the active and inactive conformations of the Epo receptor.

  1. Inflammatory cytokine tumor necrosis factor α suppresses neuroprotective endogenous erythropoietin from astrocytes mediated by hypoxia-inducible factor-2α.

    PubMed

    Nagaya, Yoshiaki; Aoyama, Mineyoshi; Tamura, Tetsuya; Kakita, Hiroki; Kato, Shin; Hida, Hideki; Saitoh, Shinji; Asai, Kiyofumi

    2014-12-01

    Interest in erythropoietin (EPO) as a neuroprotective mediator has grown since it was found that systemically administered EPO is protective in several animal models of disease. However, given that the blood-brain barrier limits EPO entry into the brain, alternative approaches that induce endogenous EPO production in the brain may be more effective clinically and associated with fewer untoward side-effects. Astrocytes are the main source of EPO in the central nervous system. In the present study we investigated the effect of the inflammatory cytokine tumor necrosis factor α (TNFα) on hypoxia-induced upregulation of EPO in rat brain. Hypoxia significantly increased EPO mRNA expression in the brain and kidney, and this increase was suppressed by TNFα in vivo. In cultured astrocytes exposed to hypoxic conditions for 6 and 12 h, TNFα suppressed the hypoxia-induced increase in EPO mRNA expression in a concentration-dependent manner. TNFα inhibition of hypoxia-induced EPO expression was mediated primarily by hypoxia-inducible factor (HIF)-2α rather than HIF-1α. The effects of TNFα in reducing hypoxia-induced upregulation of EPO mRNA expression probably involve destabilization of HIF-2α, which is regulated by the nuclear factor (NF)-κB signaling pathway. TNFα treatment attenuated the protective effects of astrocytes on neurons under hypoxic conditions via EPO signaling. The effective blockade of TNFα signaling may contribute to the maintenance of the neuroprotective effects of EPO even under hypoxic conditions with an inflammatory response. PMID:25283246

  2. Erythropoietin-mediated protection of insect brain neurons involves JAK and STAT but not PI3K transduction pathways.

    PubMed

    Miljus, N; Heibeck, S; Jarrar, M; Micke, M; Ostrowski, D; Ehrenreich, H; Heinrich, R

    2014-01-31

    The cytokine erythropoietin (Epo) initiates adaptive cellular responses to both moderate environmental challenges and tissue damaging insults in various non-hematopoietic mammalian tissues including the nervous system. Neuroprotective and neuroregenerative functions of Epo in mammals are mediated through receptor-associated Janus kinase 2 and intracellular signaling cascades that modify the transcription of Epo-regulated genes. Signal transducers and activators of transcription (STAT) and phosphoinositol-3-kinase (PI3K) represent key components of two important Epo-induced transduction pathways. Our previous study on insects revealed neuroprotective and regenerative functions of recombinant human Epo (rhEpo) similar to those in mammalian nervous tissues. Here we demonstrate that rhEpo effectively rescues primary cultured locust brain neurons from apoptotic cell death induced by hypoxia or the chemical compound H-7. The Janus kinase inhibitor AG-490 and the STAT inhibitor sc-355797 abolished protective effects of rhEpo on locust brain neurons. In contrast, inhibition of PI3K with LY294002 had no effect on rhEpo-mediated neuroprotection. The results indicate that rhEpo mediates the protection of locust brain neurons through interference with apoptotic pathways by the activation of a Janus kinase-associated receptor and STAT transcription factor(s). The involvement of similar transduction pathways in mammals and insects for the mediation of neuroprotection and support of neural regeneration by Epo indicates that an Epo/Epo receptor-like signaling system with high structural and functional similarity exists in both groups of animals. Epo-like signaling involved in tissue protection appears to be an ancient beneficial function shared by vertebrates and invertebrates. PMID:24269933

  3. Erythropoietin Ameliorates Neonatal Hypoxia-Ischemia-Induced Neurobehavioral Deficits, Neuroinflammation, and Hippocampal Injury in the Juvenile Rat.

    PubMed

    Lan, Kuo-Mao; Tien, Lu-Tai; Cai, Zhengwei; Lin, Shuying; Pang, Yi; Tanaka, Sachiko; Rhodes, Philip G; Bhatt, Abhay J; Savich, Renate D; Fan, Lir-Wan

    2016-01-01

    The hematopoietic growth factor erythropoietin (EPO) has been shown to be neuroprotective against hypoxia-ischemia (HI) in Postnatal Day 7 (P7)-P10 or adult animal models. The current study was aimed to determine whether EPO also provides long-lasting neuroprotection against HI in P5 rats, which is relevant to immature human infants. Sprague-Dawley rats at P5 were subjected to right common carotid artery ligation followed by an exposure to 6% oxygen with balanced nitrogen for 1.5 h. Human recombinant EPO (rEPO, at a dose of 5 units/g) was administered intraperitoneally one hour before or immediately after insult, followed by additional injections at 24 and 48 h post-insult. The control rats were injected with normal saline following HI. Neurobehavioral tests were performed on P8 and P20, and brain injury was examined on P21. HI insult significantly impaired neurobehavioral performance including sensorimotor, locomotor activity and cognitive ability on the P8 and P20 rats. HI insult also resulted in brain inflammation (as indicated by microglia activation) and neuronal death (as indicated by Jade B positive staining) in the white matter, striatum, cortex, and hippocampal areas of the P21 rat. Both pre- and post-treatment with rEPO significantly improved neurobehavioral performance and protected against the HI-induced neuronal death, microglia activation (OX42+) as well as loss of mature oligodendrocytes (APC-CC1+) and hippocampal neurons (Nissl+). The long-lasting protective effects of rEPO in the neonatal rat HI model suggest that to exert neurotrophic activity in the brain might be an effective approach for therapeutic treatment of neonatal brain injury induced by hypoxia-ischemia. PMID:26927081

  4. Erythropoietin activates two distinct signaling pathways required for the initiation and the elongation of c-myc

    NASA Technical Reports Server (NTRS)

    Chen, C.; Sytkowski, A. J.

    2001-01-01

    Erythropoietin (Epo) stimulation of erythroid cells results in the activation of several kinases and a rapid induction of c-myc expression. Protein kinase C is necessary for Epo up-regulation of c-myc by promoting elongation at the 3'-end of exon 1. PKCepsilon mediates this signal. We now show that Epo triggers two signaling pathways to c-myc. Epo rapidly up-regulated Myc protein in BaF3-EpoR cells. The phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 blocked Myc up-regulation in a concentration-dependent manner but had no effect on the Epo-induced phosphorylation of ERK1 and ERK2. LY294002 also had no effect on Epo up-regulation of c-fos. MEK1 inhibitor PD98059 blocked both the c-myc and the c-fos responses to Epo. PD98059 and the PKC inhibitor H7 also blocked the phosphorylation of ERK1 and ERK2. PD98059 but not LY294002 inhibited Epo induction of ERK1 and ERK2 phosphorylation in normal erythroid cells. LY294002 blocked transcription of c-myc at exon 1. PD98059 had no effect on transcription from exon 1 but, rather, blocked Epo-induced c-myc elongation at the 3'-end of exon 1. These results identify two Epo signaling pathways to c-myc, one of which is PI3K-dependent operating on transcriptional initiation, whereas the other is mitogen-activated protein kinase-dependent operating on elongation.

  5. Erythropoietin has an anti-myeloma effect - a hypothesis based on a clinical observation supported by animal studies.

    PubMed

    Mittelman, Moshe; Zeidman, Aliza; Kanter, Pazit; Katz, Odelia; Oster, Howard; Rund, Debbora; Neumann, Drorit

    2004-03-01

    Recombinant human erythropoietin (rHuEpo) was introduced into clinical practice more than a decade ago, and has been found to be effective in the treatment of several types of anemia, including anemia of end-stage renal failure and cancer-related anemia. No study has suggested that Epo might have an effect on the biology of the disease, nor any survival advantage to cancer patients treated with Epo for anemia has been reported. Here we report six patients with advanced multiple myeloma (MM) with very poor prognostic features, whose expected survival was <6 months. All six patients were treated with rHuEpo for their anemia, either without any chemotherapy or very mild chemotherapy for a short time. Yet, surprisingly they lived for 45-133 months totally from MM diagnosis and 38-94 months with rHuEpo (with a good quality of life). In fact, one patient, is still alive and well, more than 8 yr after chemotherapy was discontinued because of a resistant aggressive disease. The course in these six MM patients led us to hypothesize that Epo might have an antineoplastic or antimyeloma effect. We proceeded and tested that hypothesis in mouse models of myeloma (Mittelman M et al., Proc Natl Acad Sci USA 98:5181,2001). In these models we confirmed that rHuEpo induced tumor regression in about 50% of the BALB/c mice inoculated with MOPC-315 myeloma cells. We then presented evidence that the mechanism is a new immune-mediated phenomenon, via activation of CD8+ T cells. Furthermore, evidence from the literature supports the antineoplastic effect of Epo. Epo might be used as an adjunct immune treatment in various malignant diseases, in addition to the current regimens and chemotherapeutic protocols. Future trials should determine the role of Epo in myeloma and cancer treatment, besides clarifying concerns about the presence of Epo receptors on some tumor cells.

  6. Erythropoietin Receptor Antagonist Suppressed Ectopic Hemoglobin Synthesis in Xenografts of HeLa Cells to Promote Their Destruction.

    PubMed

    Yasuda, Yoshiko; Fujita, Mitsugu; Koike, Eiji; Obata, Koshiro; Shiota, Mitsuru; Kotani, Yasushi; Musha, Terunaga; Tsuji-Kawahara, Sachiyo; Satou, Takao; Masuda, Seiji; Okano, Junko; Yamasaki, Harufumi; Okumoto, Katsumi; Uesugi, Tadao; Nakao, Shinichi; Hoshiai, Hiroshi; Mandai, Masaki

    2015-01-01

    The aim of this study is to explore a cause-oriented therapy for patients with uterine cervical cancer that expresses erythropoietin (Epo) and its receptor (EpoR). Epo, by binding to EpoR, stimulates the proliferation and differentiation of erythroid progenitor cells into hemoglobin-containing red blood cells. In this study, we report that the HeLa cells in the xenografts expressed ε, γ, and α globins as well as myoglobin (Mb) to produce tetrameric α2ε2 and α2γ2 and monomeric Mb, most of which were significantly suppressed with an EpoR antagonist EMP9. Western blotting revealed that the EMP9 treatment inhibited the AKT-pAKT, MAPKs-pMAPKs, and STAT5-pSTAT5 signaling pathways. Moreover, the treatment induced apoptosis and suppression of the growth and inhibited the survival through disruption of the harmonized hemoprotein syntheses in the tumor cells concomitant with destruction of vascular nets in the xenografts. Furthermore, macrophages and natural killer (NK) cells with intense HIF-1α expression recruited significantly more in the degenerating foci of the xenografts. These findings were associated with the enhanced expressions of nNOS in the tumor cells and iNOS in macrophages and NK cells in the tumor sites. The treated tumor cells exhibited a substantial number of perforations on the cell surface, which indicates that the tumors were damaged by both the nNOS-induced nitric oxide (NO) production in the tumor cells as well as the iNOS-induced NO production in the innate immune cells. Taken together, these data suggest that HeLa cells constitutively acquire ε, γ and Mb synthetic capacity for their survival. Therefore, EMP9 treatment might be a cause-oriented and effective therapy for patients with squamous cell carcinoma of the uterine cervix. PMID:25874769

  7. The Influence of Artificially Introduced N-Glycosylation Sites on the In Vitro Activity of Xenopus laevis Erythropoietin

    PubMed Central

    Nagasawa, Kazumichi; Meguro, Mizue; Sato, Kei; Tanizaki, Yuta; Nogawa-Kosaka, Nami; Kato, Takashi

    2015-01-01

    Erythropoietin (EPO), the primary regulator of erythropoiesis, is a heavily glycosylated protein found in humans and several other mammals. Intriguingly, we have previously found that EPO in Xenopus laevis (xlEPO) has no N-glycosylation sites, and cross-reacts with the human EPO (huEPO) receptor despite low homology with huEPO. In this study, we introduced N-glycosylation sites into wild-type xlEPO at the positions homologous to those in huEPO, and tested whether the glycosylated mutein retained its biological activity. Seven xlEPO muteins, containing 1–3 additional N-linked carbohydrates at positions 24, 38, and/or 83, were expressed in COS-1 cells. The muteins exhibited lower secretion efficiency, higher hydrophilicity, and stronger acidic properties than the wild type. All muteins stimulated the proliferation of both cell lines, xlEPO receptor-expressing xlEPOR-FDC/P2 cells and huEPO receptor-expressing UT-7/EPO cells, in a dose-dependent manner. Thus, the muteins retained their in vitro biological activities. The maximum effect on xlEPOR-FDC/P2 proliferation was decreased by the addition of N-linked carbohydrates, but that on UT-7/EPO proliferation was not changed, indicating that the muteins act as partial agonists to the xlEPO receptor, and near-full agonists to the huEPO receptor. Hence, the EPO-EPOR binding site in X. laevis locates the distal region of artificially introduced three N-glycosylation sites, demonstrating that the vital conformation to exert biological activity is conserved between humans and X. laevis, despite the low similarity in primary structures of EPO and EPOR. PMID:25898205

  8. Effects of erythropoietin preconditioning on rat cerebral ischemia-reperfusion injury and the GLT-1/GLAST pathway

    PubMed Central

    YU, DAIHUA; FAN, YUANHUA; SUN, XUDE; YAO, LINONG; CHAI, WEI

    2016-01-01

    The aim of the present study was to investigate whether erythropoietin (EPO) preconditioning affects the expression of glutamate transporter 1 (GLT-1) and glutamate aspartate transporter (GLAST) and protects against rat cerebral ischemia-reperfusion injury. A total of 140 Sprague Dawley rats were randomly assigned to one of the following four groups: Sham, EPO-sham, middle cerebral artery occlusion (MCAO) and EPO-MCAO. Neurological function scores were obtained 24, 36 and 72 h after reperfusion. Seventy-two hours after the induction of cerebral ischemia-reperfusion, the number of apoptotic neural cells and the cerebral infarct volume of each group were measured. The mRNA levels of GLT-1 and GLAST were determined using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis, while the GLT-1 and GLAST protein levels were assessed using western blotting. The cerebral infarct volume was significantly increased in the MCAO group compared with that in the sham group (P<0.01); however, the infarct volume of the EPO-MCAO group was significantly lower than that of the MCAO group (P<0.01). In addition, the number of apoptotic cells found in the MCAO group was higher than that in the sham group (P<0.01), but the number of apoptotic cells in the EPO-MCAO group was significantly lower than that in the MCAO group (P<0.01). The GLT-1 and GLAST mRNA and protein levels were significantly decreased 72 h after the cerebral ischemia-reperfusion (P<0.01) compared with those in the sham group, whereas the same levels were increased significantly in the EPO-MCAO group relative to those in the MCAO group (P<0.01). In conclusion, EPO preconditioning protected against cerebral ischemia-reperfusion injury and upregulated the GLT-1 and GLAST expression. PMID:26893639

  9. Effect of androgen therapy and anemia on serum erythropoietin levels in patients with aplastic anemia and myelodysplastic syndromes.

    PubMed

    Piedras, J; Hernández, G; López-Karpovitch, X

    1998-02-01

    Immunoreactive serum erythropoietin (EPO) was measured in anemic and non-anemic patients with acquired non-severe aplastic anemia (AA; n = 22) and myelodysplastic syndromes (MDS; n = 31) receiving or not androgens to examine the effect of androgen therapy and anemia on EPO levels in these disorders. Soluble transferrin receptor (TfR) and absolute reticulocyte count (ARC) were also assayed in order to evaluate erythropoietic activity. AA and MDS patients were stratified for anemia and androgen treatment as follows: 12 untreated anemic patients; 17 anemic patients during androgen therapy; 14 non-anemic patients without any treatment (> 1 year); and 10 non-anemic patients on androgen therapy. Although EPO levels in non-anemic patients were significantly higher than in healthy controls (n = 29) no statistically significant differences in Hb and EPO values were found between non-anemic patients receiving or not androgen therapy. In the linear regression analysis between Hb and log EPO concentration, no statistically significant differences in the slopes between untreated and androgen-treated anemic groups nor between both groups and patients with iron deficiency anemia (n = 23) were observed. However, the y intercept (log EPO) of regression line was significantly higher in androgen-treated anemic patients than in the androgen therapy-free anemic group. Serum TfR levels were higher in treated than in untreated anemic patients, whereas ARC was not different between both groups. These data seemingly indicate that (1) androgens at pharmacological doses do not increase serum EPO levels in non-anemic AA and MDS patients, and (2) in patients with AA and MDS, androgen-driven EPO stimulation is appreciably enhanced by anemia.

  10. Ventilatory response to acute hypoxia in transgenic mice over-expressing erythropoietin: effect of acclimation to 3-week hypobaric hypoxia.

    PubMed

    Villafuerte, Francisco C; Cárdenas-Alayza, Rosa; Macarlupú, José Luis; Monge-C, Carlos; León-Velarde, Fabiola

    2007-09-30

    We used transgenic mice constitutively over-expressing erythropoietin ("tg6" mice) and wild-type (wt) mice to investigate whether the high hematocrit (hct), consequence of Epo over-expression affected: (1) the normoxic ventilation (V (E)) and the acute hypoxic ventilatory response (HVR) and decline (HVD), (2) the increase in ventilation observed after chronic exposure to hypobaric hypoxia (430mmHg for 21 days), (3) the respiratory "blunting", and (4) the erythrocythemic response induced by chronic hypoxia exposure. V (E) was found to be similar in tg6 and wt mice in normoxia (FIO2=0.21). Post-acclimation V (E) was significantly elevated in every time point in wt mice at FIO2=0.10 when compared to pre-acclimation values. In contrast, tg6 mice exhibited a non-significant increase in V (E) throughout acute hypoxia exposure. Changes in V (E) are associated with adjustments in tidal volume (V(T)). HVR and HVD were independent of EE in tg6 and wt mice before chornic hypoxia exposure. HVR was significantly greater in wt than in tg6 mice after chronic hypoxia. After acclimation, HVD decreased in tg6 mice. Chronic hypoxia exposure caused hct to increase significantly in wt mice, while only a marginal increase occurred in the tg6 group. Although pre-existent EE does not appear to have an effect on HVR, the observation of alterations on V(T) suggests that it may contribute to time-dependent changes in ventilation and in the acute HVR during exposure to chronic hypoxia. In addition, our results suggest that EE may lead to an early "blunting" of the ventilatory response.

  11. Erythropoietin loaded microspheres prepared from biodegradable LPLG-PEO-LPLG triblock copolymers: protein stabilization and in-vitro release properties.

    PubMed

    Morlock, M; Kissel, T; Li, Y X; Koll, H; Winter, G

    1998-12-01

    Biodegradable microspheres containing recombinant human Erythropoietin (EPO) were prepared from ABA triblock copolymers, consisting of hydrophobic poly(l-lactic-co-glycolic acid) A blocks and hydrophilic polyethylenoxide (PEO) B blocks. Different polymer compositions were studied for the microencapsulation of EPO using a modified double-emulsion process (W/O/W). The encapsulation efficiency for EPO, ranging from 72% to 99% was quite acceptable. The formation of high molecular weight EPO aggregates, however, was higher than in poly(d,l-lactide-co-glycolide) (PLG) microparticles. Using different excipients with known protein stabilizing properties, such as Bovine Serum Albumin (BSA), Poly-l-Histidine (PH), Poly-l-Arginine (PA) or a combination of PA with Dextran 40 (D40), the EPO aggregate content was significantly reduced to <5% of the encapsulated EPO. In contrast to PLG, ABA triblockcopolymers containing >7 mol % PEO, allowed a continuous release of EPO from microspheres for up to 2 weeks under in-vitro conditions. The release profile was comparable to FITC-Dextran 40 kDa (FD 40) loaded microspheres in the initial release phase, while EPO release was leveling off at later time points. BSA additionally prolonged the EPO release, while blends of PLG and PEO did not generate continuous EPO release profiles. LPLG-PEO-LPLG triblock-copolymers (35 mol % PEO; 30 kDa) in combination with 5% BSA yielded both an acceptable level of EPO aggregates and a continuous release profile under in-vitro conditions for up to 2 weeks. The formation of EPO aggregates at later time points is probably induced by acidic cleavage products of the biodegradable polymer and requires further optimization of the ABA polymer composition.

  12. Friend spleen focus-forming virus induces factor independence in an erythropoietin-dependent erythroleukemia cell line.

    PubMed Central

    Ruscetti, S K; Janesch, N J; Chakraborti, A; Sawyer, S T; Hankins, W D

    1990-01-01

    Erythroid cells from mice infected with the polycythemia-inducing strain of Friend spleen focus-forming virus (SFFVP), unlike normal erythroid cells, can proliferate and differentiate in apparent absence of the erythroid hormone erythropoietin (Epo). The unique envelope glycoprotein encoded by SFFV has been shown to be responsible for this biological effect. The recent isolation of an Epo-dependent erythroleukemia cell line, HCD-57, derived from a mouse infected at birth with Friend murine leukemia virus, afforded us the opportunity to study the direct effect of SFFVP on a homogeneous population of factor-dependent cells. The introduction of SFFVP in complex with various helper viruses into these Epo-dependent cells efficiently and reproducibly gave rise to lines which expressed high levels of SFFV and were factor independent. SFFV appears to be unique in its ability to abrogate the factor dependence of Epo-dependent HCD-57 cells, since infection of these cells with retroviruses carrying a variety of different oncogenes had no effect. The induction of Epo independence by SFFV does not appear to involve a classical autocrine mechanism, since there is no evidence that the factor-independent cells synthesize or secrete Epo or depend on it for their growth. However, the SFFV-infected, factor-independent cells had significantly fewer receptors available for binding Epo than their factor-dependent counterparts had, raising the possibility that the induction of factor independence by the virus may be due to the interaction of an SFFV-encoded protein with the Epo receptor. Images PMID:2154592

  13. Analysis of the correlation between serum resistin and the variability of erythropoietin responsiveness in patients with chronic kidney disease

    PubMed Central

    ZHANG, HONGHAO; LI, XIUJIANG; KAN, YANHONG; YANG, FAN; HOU, YUE; DU, YUJUN

    2015-01-01

    Chronic kidney disease (CKD) is commonly accompanied by inflammation and anemia; however, the pathogenesis of CKD is unclear. Expression of resistin, a cysteine-rich secretory plasma protein, is correlated with the expression of tumor necrosis factor (TNF)-α, interleukin (IL)-6 and lipoprotein-associated phospholipase A2, indicating that resistin may be involved in inflammatory events. In addition, inflammation inhibits the activity of erythropoietin (EPO) and, thus, erythropoiesis. The aim of the present study was to analyze the correlation between serum resistin and the variability of EPO responsiveness in CKD patients. The levels of serum creatinine (SCr), C-reactive protein (CRP), total cholesterol, triglycerides, IL-6 and serum resistin were measured in the samples obtained from 138 CKD patients and healthy control subjects. The levels of serum resistin in the CKD groups with and without hemodialysis were significantly higher than those observed in the normal control group (P<0.01) and the levels of serum resistin in the hemodialysis CKD group were higher than those observed in the CKD group without dialysis (P<0.01). The levels of serum resistin in patients in the randomly selected CKD group (with hemodialysis) were positively correlated with the duration of dialysis and the levels of SCr and CRP (P<0.05), however, were negatively correlated with the estimated glomerular filtration rate. The EPO resistance index (ERI) was identified to be associated with body mass index and the levels of CRP and resistin; furthermore, EPO reactivity was correlated with the level of resistin and ERI. The levels of serum resistin were correlated with the variability in EPO responsiveness that was observed in the CKD patients. Therefore, the measurement of serum resistin may aid with understanding the mechanisms, clinical diagnosis and treatment of CKD. PMID:26640574

  14. Erythropoietin inhibits gluconeogenesis and inflammation in the liver and improves glucose intolerance in high-fat diet-fed mice.

    PubMed

    Meng, Ran; Zhu, Dalong; Bi, Yan; Yang, Donghui; Wang, Yaping

    2013-01-01

    Erythropoietin (EPO) has multiple biological functions, including the modulation of glucose metabolism. However, the mechanisms underlying the action of EPO are still obscure. This study is aimed at investigating the potential mechanisms by which EPO improves glucose tolerance in an animal model of type 2 diabetes. Male C57BL/6 mice were fed with high-fat diet (HFD) for 12 weeks and then treated with EPO (HFD-EPO) or vehicle saline (HFD-Con) for two week. The levels of fasting blood glucose, serum insulin and glucose tolerance were measured and the relative levels of insulin-related phosphatidylinositol 3-kinase (PI3K)/Akt, insulin receptor (IR) and IR substrate 1 (IRS1) phosphorylation were determined. The levels of phosphoenolpyruvate carboxykinase (PEPCK), glucose-6- phosphatase (G6Pase), toll like receptor 4 (TLR4), tumor necrosis factor (TNF)-α and IL-6 expression and nuclear factor-κB (NF-κB) and c-Jun N-terminal kinase (JNK), extracellular-signal-regulated kinase (ERK) and p38 MAPK activation in the liver were examined. EPO treatment significantly reduced the body weights and the levels of fasting blood glucose and serum insulin and improved the HFD-induced glucose intolerance in mice. EPO treatment significantly enhanced the levels of Akt, but not IR and IRS1, phosphorylation, accompanied by inhibiting the PEPCK and G6Pase expression in the liver. Furthermore, EPO treatment mitigated the HFD-induced inflammatory TNF-α and IL-6 production, TLR4 expression, NF-κB and JNK, but not ERK and p38 MAPK, phosphorylation in the liver. Therefore, our data indicated that EPO treatment improved glucose intolerance by inhibiting gluconeogenesis and inflammation in the livers of HFD-fed mice. PMID:23326455

  15. Sialic acid-specific affinity chromatography for the separation of erythropoietin glycoforms using serotonin as a ligand.

    PubMed

    Meininger, M; Stepath, M; Hennig, R; Cajic, S; Rapp, E; Rotering, H; Wolff, M W; Reichl, U

    2016-02-15

    Recombinant human erythropoietin (rhEPO) is an important CHO cell-derived glycoprotein and the degree of sialylation of this hormone is crucial for its in vivo bioactivity. In order to improve the purification process serotonin as a potential affinity ligand was tested for preparative chromatographic separation of rhEPO glycoforms into fractions of different degrees of sialylation. Therefore, two chromatographic matrices were prepared by immobilizing serotonin on CNBr- and NHS-Sepharose™. First it was shown both matrices bind rhEPO only in its sialylated form. Results indicate that binding is pH independent between pH 3.5 to 8 suggesting it is not only based on electrostatic interactions. Second, after optimal binding conditions were identified, semi-purified rhEPO was loaded onto both matrices and eluted using a stepwise elution gradient of sodium chloride. For comparison same affinity purification experiments were performed using wheat germ agglutinin-coupled agarose, a lectin known for its affinity towards sialylated glycoproteins. To monitor changes in N-glycan fingerprint, eluate fractions were analyzed by multiplexed capillary gel electrophoresis coupled to laser-induced fluorescence (xCGE-LIF). For the serotonin matrices an increasing degree of sialylation was observed from the first to the third elution fraction while purity of rhEPO could be increased at the same time. The late elution fractions of serotonin-coupled CNBr- and NHS-Sepharose™ also showed an overall sialylation degree exceeding that of the starting material. In contrast, for rhEPO bound to wheat germ agglutinin-coupled agarose, no distinct change in the degree of sialylation could be observed after elution. Overall, these encouraging results highlight the potential of serotonin as a chromatographic ligand for the improvement of pharmaceutical purification processes of rhEPO. PMID:26851523

  16. Thoughts for New Administrators

    ERIC Educational Resources Information Center

    LaCrosse, E. Robert

    1977-01-01

    Discusses issues in program administration, with specific reference to preschool programs. Some of the issues discussed include coping with conflict, leadership, the sharing of power, and honesty. A short annotated bibliography is included. (BD)

  17. Evaluation of Administrators.

    ERIC Educational Resources Information Center

    Ryan, Edmund G.

    1979-01-01

    Guidelines are presented for the evaluation of college administrators: (1) purposes of the evaluation; (2) approaches to evaluation; (3) criteria for evaluation; (4) participants or evaluators in the process; and (5) evaluation results and use of results. (GDC)

  18. Serving the Space Administration

    ERIC Educational Resources Information Center

    Campbell, Jack E.; Thompson, Arthur W.

    1974-01-01

    The purpose of the current program was to establish an upward mobility program that afforded employees an opportunity to improve their credibility in job opportunity selection under the directives of the National Aeronautics and Space Administration. (Author/RK)

  19. Goldstone (GDSCC) administrative computing

    NASA Technical Reports Server (NTRS)

    Martin, H.

    1981-01-01

    The GDSCC Data Processing Unit provides various administrative computing services for Goldstone. Those activities, including finance, manpower and station utilization, deep-space station scheduling and engineering change order (ECO) control are discussed.

  20. Administrators Speak Out

    ERIC Educational Resources Information Center

    Miner, Fred V.

    1976-01-01

    A summary is presented of the issues and concerns discussed and the recommendations made by the American Vocational Association's (AVA) Administration Policy Committee during its meeting at the 1975 AVA Convention. (AJ)