Administration Of Recombinant Human Erythropoietin Research Articles

Short-term administration of recombinant human erythropoietin decreases B cell number in human peripheral blood

ObjectivesThere are conflicting results on the influence of recombinant human erythropoietin (rHuEPO) administration to lymphocytes, especially to B cells. MethodsWe analyzed peripheral white blood cell (WBC) subsets in patients who received one bolus administration of rHuEPO. 119 autologous blood donors were enrolled in this study. Fourty-nine out of them were treated with rHuEPO. Blood samples were obtained before the first phlebotomy and one week later before the second one. By flow cytometry, we measured the numbers of WBC, lymphocytes, dendritic cells, CD4+ T cells, CD8+ T cells, natural killer (NK) cells, B cells, monocytes, and neutrophils, further details of B cell subsets. ResultsIn the EPO-treatment group, absolute numbers of lymphocytes, especially CD8+ T cells, NK cells, and B cells, significantly decreased after rHuEPO administration. In B cell subsets, absolute numbers of naïve B cells and IgD−CD27− B cells significantly decreased. Other B cell subsets, such as transitional B cells, memory B cells, and marginal zone B cells, also showed a decreasing trend. ConclusionThese findings suggest that a single administration of rHuEPO can influence human immune system via reduction of B cell number in peripheral blood.

Validation of whole-blood transcriptome signature during microdose recombinant human erythropoietin (rHuEpo) administration

BackgroundRecombinant human erythropoietin (rHuEpo) can improve human performance and is therefore frequently abused by athletes. As a result, the World Anti-Doping Agency (WADA) introduced the Athlete Biological Passport (ABP) as an indirect method to detect blood doping. Despite this progress, challenges remain to detect blood manipulations such as the use of microdoses of rHuEpo.MethodsForty-five whole-blood transcriptional markers of rHuEpo previously derived from a high-dose rHuEpo administration trial were used to assess whether microdoses of rHuEpo could be detected in 14 trained subjects and whether these markers may be confounded by exercise (n = 14 trained subjects) and altitude training (n = 21 elite runners and n = 4 elite rowers, respectively). Differential gene expression analysis was carried out following normalisation and significance declared following application of a 5% false discovery rate (FDR) and a 1.5 fold-change. Adaptive model analysis was also applied to incorporate these markers for the detection of rHuEpo.ResultsALAS2, BCL2L1, DCAF12, EPB42, GMPR, SELENBP1, SLC4A1, TMOD1 and TRIM58 were differentially expressed during and throughout the post phase of microdose rHuEpo administration. The CD247 and TRIM58 genes were significantly up- and down-regulated, respectively, immediately following exercise when compared with the baseline both before and after rHuEpo/placebo. No significant gene expression changes were found 30 min after exercise in either rHuEpo or placebo groups. ALAS2, BCL2L1, DCAF12, SLC4A1, TMOD1 and TRIM58 tended to be significantly expressed in the elite runners ten days after arriving at altitude and one week after returning from altitude (FDR > 0.059, fold-change varying from 1.39 to 1.63). Following application of the adaptive model, 15 genes showed a high sensitivity (≥ 93%) and specificity (≥ 71%), with BCL2L1 and CSDA having the highest sensitivity (93%) and specificity (93%).ConclusionsCurrent results provide further evidence that transcriptional biomarkers can strengthen the ABP approach by significantly prolonging the detection window and improving the sensitivity and specificity of blood doping detection. Further studies are required to confirm, and if necessary, integrate the confounding effects of altitude training on blood doping.

Decrease in Oxidative Stress Parameters after Post-Ischaemic Recombinant Human Erythropoietin Administration in the Hippocampus of Rats Exposed to Focal Cerebral Ischaemia.

Recombinant human erythropoietin (rhEpo) is a multi-functional drug with antioxidant potential. However, the underlying molecular mechanisms of its action are still unclear. The purpose of this study was to investigate the effects of rhEpo on the brain infarct volume as well as on the levels of the neuronal damage, oxidative stress parameters and active caspase-3, nuclear factor erythroid 2-related factor 2 (Nrf2) and haemeoxygenase-1 (HO-1) expressions in the hippocampi of rats exposed to the right middle cerebral artery occlusion (MCAO) for 1hr. Ischaemic animals received either vehicle or rhEpo (5000IU/kg, i.p.) immediately or 3hr after the induction of ischaemia. Sham-operated, vehicle-treated animals served as the control group. Rats were killed 24hr after the onset of the ischaemic or sham experimental procedure. MCAO caused ipsilateral brain infarction within the striatum and cortex. In the CA1 region of the hippocampi, we did not find significant neuronal loss, but a statistically significant rise in the active caspase-3 and Nrf2 protein expressions was registered. We detected also significant increases in the hippocampal levels of oxidative stress parameters (thiobarbituric acid-reactive substances, superoxide dismutase, glutathione peroxidase). Post-ischaemic administration of rhEpo significantly reduced the brain infarct volume, decreased levels of all tested oxidative stress parameters and increased the Nrf2 expression level. These findings suggest that decrease in oxidative stress parameters in the hippocampus could be an early indicator of post-ischaemic neuroprotective effect of rhEpo in rats exposed to focal cerebral ischaemia and that this effect could be attributable to additional post-ischaemic activation of Nrf2 endogenous antioxidant system.

Recovery from experimental parkinsonism by intrastriatal application of erythropoietin or EPO-releasing neural precursors

An extensive literature has shown a powerful neuroprotective action of Erythropoietin (EPO) both in vivo and in vitro. This study shows that EPO, whether ectopically administered or released by neural precursors, does reverse MPTP-induced parkinsonism in mice. Unilateral stereotaxic injection of 2.5 × 105 erythropoietin-releasing neural precursor cells (Er-NPCs) rescued degenerating striatal dopaminergic neurons and promoted behavioral recovery as shown by three independent behavioral tests. These effects were replicated through direct intrastriatal administration of recombinant human EPO. At the end of the observational period, most of the transplanted Er-NPCs were vital and migrated via the striatum to reach Substantia Nigra. The restorative effects appear to be mediated by EPO since co-injection of anti-EPO or anti-EPOR antibodies antagonized the positive outcomes. Furthermore, this report supports the neuroprotective action of EPO, which may also be achieved via administration of EPO-releasing cells such as Er-NPCs.

One-Time Administration of Recombinant Human Erythropoietin Decreases B Cell in Human Peripheral Blood

Introduction:Erythropoietin (EPO) is hematopoietic factors participating in red blood cell production by accelerating cell proliferation and inhibition of apoptosis of erythroblast. It is reported that EPO has pleiotropic effects including anti-apoptotic action for some cells, antioxidant action, vascularization action, and promoting cell division in addition to stimulation of erythropoiesis as well, whereas there are conflicting results of small cohorts as to its effect on blood immune cells. We analyzed peripheral white blood cell subsets in patients who received one bolus administration of recombinant human erythropoietin (rHuEPO) to examine the effect of EPO on human immune system.Materials and Methods:One hundred twenty eight autologous blood donors (male/female 64/64) in Gunma University Hospital were enrolled in this study after written informed consent. All the patients had no infections or inflammation. Fifty six patients were administered rHuEPO (Epoetin alpha or Epoetin beta, 24,000 IU) once after blood donation because of low hemoglobin concentration (EPO group) and 72 were not administered (non-EPO group). Peripheral blood samples were obtained at the time of the first phlebotomy and after 7 days from the same patient.We measured the number of WBC, lymphocytes, myeloid dendritic cells (mDC), plasmacytoid dendritic cells (pDC), CD4+ T cells, CD8+ T cells, Natural killer (NK) cells, B cells, monocytes, and neutrophils of peripheral blood before and after rHuEPO administration by flow cytometry. Also we compared EPO group with non-EPO group. Paired and unpaired Student’s t-test were used to compare absolute counts and percentages of each cell, P values < 0.05 were considered significant. This study was approved by the research ethics committee of our hospital.Results:In EPO group, absolute number and percentage of lymphocyte in WBC decreased significantly after rHuEPO administration from 1754.9 ± 535.4/µl to 1627.2 ± 467.7/µl, (p<0.01) and from 30.9 ± 8.00% to 28.2 ± 6.77% (p<0.01), respectively. The numbers of whole WBC, mDC, pDC, monocyte and neutrophil did not show significant changes. In regards to lymphocyte subset, absolute number of CD8+ T cell significantly decreased from 358.9 ± 257.0/µl to 311.5 ± 210.9/µl (p=0.04) and that of NK cell from 290.6 ± 157.6/µl to 257.4 ± 141.8/µl (p<0.01), but their percentages did not change. B cells significantly decreased in absolute number from 258.3 ± 154.2 /µl to 221.6 ± 129.2 /µl (p<0.01) and also in percentages from 14.8 ± 6.92% to 13.6 ± 6.07% (p<0.01). Regarding B cell subsets, absolute number of Naïve B cells significantly decreased from 171.3 ± 93.5 /µl to 153.0 ± 84.2 /µl (p<0.01); moreover other B cell subsets, such as transitional B cells, memory B cells and marginal zone B cells, also showed a trend of decrease, so that the percentage of Naïve B cell did not change in total B cells. These suggested that whole B cell decreased, not a specific subset of B cell. In non-EPO group, there was no significant change in any type of immune cells. When compared the rate of change between before and after rHuEPO treatment, B cell number significant decreased in EPO group compared to non-EPO group (0.87 vs 0.98, p<0.01). Multiple regression analysis revealed rHuEPO administration significantly influenced B cell number (p=0.015); however age, sex, disease status and the type of rHuEPO preparations (epoetin alfa or beta) did not influence.Conclusion:These findings suggested that just one administration of rHuEPO influenced human immune system, especially via reduction of B cell in peripheral blood. DisclosuresNo relevant conflicts of interest to declare.

Delayed administration of recombinant human erythropoietin reduces apoptosis andinflammation and promotes myelin repair and functional recovery following spinal cord compressive injury in rats.

A previous study showed that a 1-h delay in treatment of thoracic spinal cord injury (SCI) with recombinant human erythropoietin (rhEPO) lacked neuroprotective efficacy. The aim of the present study was to reassess delayed administration of different doses of rhEPO on acute spinal cord compressive injury in rats. The experiment was divided into first and second stages, which SCI rats were observed for 4 and 28 days, respectively. All rats were randomly divided into four groups at both stages: control group, and rhEPO-3,000U (Unit), rhEPO-4,000U and rhEPO-5,000U groups. SCI rats received rhEPO treatment at different time points. The primary indicators were locomotor recovery, histopathology, apoptotic index, inflammatory index, ultrastructural scoring system and volume of areas of demyelination. The most significant locomotor functional and histopathological improvements and the best myelin protection were observed after administration of 5,000 U/kg rhEPO. rhEPO at 3,000, 4,000 and 5,000 U/kg showed similar ultrastructural neuroprotection, as well as similar inhibition of apoptosis and regulation of inflammation. Delayed administration of rhEPO can reduce apoptosis and inflammation, and promote myelin repair and functional recovery following spinal cord compressive injury in rats.

Erythropoietin corrects anaemia and reduces the risk of blood transfusion in people with chronic kidney disease, but has uncertain effects on other patient-level outcomes.

Commentary on: Cody JD, Hodson EM. Recombinant human erythropoietin versus placebo or no treatment for the anaemia of chronic kidney disease in people not requiring dialysis. Cochrane Database Syst Rev 2016;(1):CD003266 Anaemia frequency and severity worsen with advancing chronic kidney disease (CKD) and are associated with quality-of-life (QOL) impairment, morbidity and mortality.1 Deficient renal erythropoietin production is a major cause and can be corrected by recombinant human erythropoietin (rhEPO) administration.1 This may improve clinical outcomes, including delaying dialysis. Conversely, rhEPO therapy causes hypertension,2 thereby potentially accelerating CKD progression. This systematic review and meta-analysis examined rhEPO effects on clinical outcomes in anaemic, pre-dialysis CKD patients. Randomised controlled trials (RCTs) comparing rhEPO treatment with either placebo …

Impacts of Erythropoietin on Vascular Endothelial Growth Factor Receptor 2 by the Extracellular Signal-regulated Kinase Signaling Pathway in a Neonatal Rat Model of Periventricular White Matter Damage

To explore the impacts of erythropoietin on vascular endothelial growth factor receptor 2 (VEGFR2) by the extracellular signal-regulated kinase (ERK) signaling pathway in a neonatal rat model of periventricular white matter damage. All of postnatal day 4 rats were randomized into three groups: the sham group [without hypoxia-ischemia (HI)], the HI group (HI with saline administration), and the erythropoietin (EPO) group [HI with recombinant human erythropoietin (rh-EPO) administration]. Rat pups underwent permanent ligation of the right common carotid artery, followed by 6% O2 for 2 hours or sham operation and normoxic exposure. Immediately after the HI, rats received a single intraventricular injection of rh-EPO (0.6 IU/g body mass) or saline. ERK and phosphorylation-ERK were examined at 60 minutes and 90 minutes after operation, and VEGFR2 were detected at 2 and 4 days after operation by using Western blot. At 60 minutes and 90 minutes after operation, the proteins of phosphorylation-ERK were significantly higher in HI rats than in the sham rats and significantly higher in HI+EPO rats than in the HI rats (P<0.05). Two days after operation, VEGFR2 was not significantly different between sham and HI rats. However, the proteins of VEGFR2 were increased after administration of rh-EPO (P<0.05). Four days after operation, the proteins of VEGFR2 were significantly higher in HI rats than in the sham rats and significantly higher in HI+EPO rats than in the HI rats (P<0.05). EPO may regulate VEGFR2 expression by affecting the intracranial ERK signaling pathways.

Ocular Erythropoietin Penetration after Subconjunctival Administration in Glaucomatous Rats

Purpose: The present study aimed to determine whether the subconjunctival administration of recombinant human erythropoietin (rHuEPO) reached the retina in glaucoma conditions. After subconjunctival rHuEPO administration, in a rat glaucoma model, erythropoietin (EPO) distribution in the rat's retina was studied by immunohistochemistry. Methods: Female Wistar Hannover albino rats (n = 15) were divided into 2 groups, control (n = 3) and treated (n = 12). The animals' unilateral glaucoma was induced by coagulation of episcleral veins, under general anaesthesia. After vein coagulation, 1,000 IU of rHuEPO were administered by the subconjunctival route to the treated group (n = 12). The control group (n = 3) received only a subconjunctival saline injection. The contralateral eye of each animal remained untouched. Treated group animals were euthanized at different time points, i.e. days 1, 3, 7 and 14. Bilateral enucleation was performed, and EPO distribution in the rat's retina was assessed by immunohistochemistry. Results: Glaucoma was confirmed by results of repeated intraocular pressure measurements over the experimental period. In the test group, EPO was identified in different neuroretinal cells, showing a stronger immunostaining signal during the first 2 time points in the retinal ganglion cell (RGC) layer. EPO protein was still present on day 14 after the subconjunctival injection. EPO was not detected in any of the control eyes or in any contralateral eye of the treated group. Conclusion: When administered subconjunctivally to glaucomatous eyes, rHuEPO reached the RGC layer and was still present at least 14 days after administration. The subconjunctival route was shown to be a promising alternative for ocular EPO delivery in glaucomatous conditions in a rat animal model.

Effects of dialyzer reuse on dialysis adequacy, anemia control, erythropoieting-stimulating agents use and phosphate level.

Almost 50 years have passed since the first time the concept of dialyzer reuse in hemodialyzed patients was introduced for primarily economic reasons [1]. Hemodialyzers are labeled by the Food and Drug Administration (FDA) and European Medicines Agency (EMA) as single use only. In recognition of the widespread practice of reuse in the United States, in 1996 the FDA required manufacturers to label dialyzers for multiple use and to back up this claim with data showing that the performance of the dialyzer is sustained over multiple uses [2]. In Poland there is also a tendency to stop the reuse of dialyzers. There are three major concerns with reuse: the risk of infection; biochemical and immunologic effects; and loss of performance with impairment in clearance and/or ultrafiltration [3].

Effects of an Injected Placebo on Endurance Running Performance.

This study aims to quantify the magnitude of the placebo effect of an injected placebo ("OxyRBX") purporting to have effects similar to those of recombinant human erythropoietin on endurance running performance in "real-world" field-based head-to-head competition settings. Fifteen endurance-trained club-level men (mean ± SD: age, 27.5 ± 6.8 yr; body mass index, 22.9 ± 2.0 kg·m) with 10-km personal-best record times of 39.3 ± 4.4 min completed a randomized cross-over study of 3-km races before and after 7-d "control" and "placebo" phases. During the placebo phase, participants self-administered subcutaneous saline injections daily, believing it to be OxyRBX, with no intervention during the control phase. At the start and end of each 7-d phase, 3-km running performance was assessed. Qualitative assessments of participants' perceptions and experiences were recorded throughout and during semistructured interviews on completion. Race time improved significantly more in response to placebo intervention (9.73 ± 1.96 s faster, P = 0.0005) than in response to control (1.82 ± 1.94 s faster, P = 0.41; Pinteraction = 0.02). In response to placebo, participants reported reductions in physical effort, increased potential motivation, and improved recovery. Beliefs and congruence between positive expectations of the effects of placebo and perceptions of physical change during training also appeared to impact on competitive performance. Compared to control, the injected placebo improved 3-km race time by 1.2%. This change is of clear sporting relevance but is smaller than the performance improvement elicited by recombinant human erythropoietin administration. Qualitative data suggest that placebo may have improved performance by both reducing perception of effort and increasing potential motivation, in accord with the psychobiological model for exercise performance, and that cognitive and noncognitive processes appear to have influenced placebo response.

Perioperative administration of high-dose recombinant human erythropoietin for delayed graft function prevention in kidney transplantation: a meta-analysis.

Delayed graft function (DGF) due to ischemia-reperfusion injury is a major early complication of kidney transplantation (KT). Recombinant human erythropoietin (rHuEPO) has been shown to exert nephroprotective action in animal models. We conducted a meta-analysis to explore the impact of rHuEPO on DGF in KT. Eligible studies comparing perioperative high-dose rHuEPO with placebo or no therapy for prevention of DGF were identified through MEDLINE, CENTRAL, and Transplant Library. Their design and data were assessed by two independent reviewers. Among 737 examined studies, four randomized controlled trials, involving 356 recipients of kidney allografts from deceased donors, fulfilled inclusion criteria. Statistical heterogeneity across studies was not significant (P = 0.98, I(2) = 0%). In a random effects model, no significant difference was found in the occurrence of DGF (odds ratio: 0,74, 95% CI: 0.47-1.18, P = 0.21). At 4 weeks after KT, the rHuEPO group exhibited higher systolic blood pressure (mean difference: 6.47 mmHg, 95% CI: 1.25-11.68, P = 0.02). Perioperative, high-dose rHuEPO administration does not prevent DGF in deceased donor KT. Furthermore, it is associated with higher systolic blood pressure leading to safety concerns. Nonerythropoietic rHuEPO derivatives, designed for nephroprotective action without increasing cardiovascular risk, might prove an alternative but still are at early stages of development.

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

Despite improved survival, many preterm infants undergo subsequent neurodevelopmental impairment. To date, no neuroprotective therapies have been implemented into clinical practice. Erythropoietin, a haematopoietic cytokine used for treatment of anaemia of prematurity, has been shown to have neuroprotective and neuroregenerative effects on the brain in many experimental studies. The aim of the study was to assess the effect of recombinant human erythropoietin on the microstructural development of the cerebral white matter using tract-based spatial statistics performed at term equivalent age. A randomized, double-blind placebo-controlled, prospective multicentre study applying recombinant human erythropoietin in the first 42 h after preterm birth entitled 'Does erythropoietin improve outcome in preterm infant' was conducted in Switzerland (NCT00413946). Preterm infants were given recombinant human erythropoietin (3000 IU) or an equivalent volume of placebo (NaCl 0.9%) intravenously before 3 h of age after birth, at 12-18 h and at 36-42 h after birth. High resolution diffusion tensor imaging was obtained at 3 T in 58 preterm infants with mean (standard deviation) gestational age at birth 29.75 (1.44) weeks, and at scanning at 41.1 (2.09) weeks. Imaging was performed at a single centre. Voxel-wise statistical analysis of the fractional anisotropy data was carried out using tract-based spatial statistics to test for differences in fractional anisotropy between infants treated with recombinant human erythropoietin and placebo using a general linear model, covarying for the gestational age at birth and the corrected gestational age at the time of the scan. Preterm infants treated with recombinant human erythropoietin demonstrated increased fractional anisotropy in the genu and splenium of the corpus callosum, the anterior and posterior limbs of the internal capsule, and the corticospinal tract bilaterally. Mean fractional anisotropy was significantly higher in preterm 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.

Individualized model discovery: The case of anemia patients

The universal sequel to chronic kidney condition (CKD) is anemia. Patients of anemia have kidneys that are incapable of performing certain basic functions such as sensing of oxygen levels to secrete erythropoietin when red blood cell counts are low. Under such conditions, external administration of human recombinant erythropoietin (EPO) is administered as alternative to improve conditions of CKD patients by increasing their hemoglobin (Hb) levels to a given therapeutic range.Presently, EPO dosing strategies extensively depend on packet inserts and on “average” responses to the medication from previous patients. Clearly dosage strategies based on these approaches are, at best, nonoptimal to EPO medication and potentially dangerous to patients that do not adhere to the notion of expected “average” response. In this work, a technique called semi-blind robust identification is provided to uniquely identify models of the individual patients of anemia based on their actual Hb responses and EPO administration. Using the a priori information and the measured input-output data of the individual patients, the procedure identifies a unique model consisting of a nominal model and the associated model uncertainty for the patients. By incorporating the effects of unknown system initial conditions, considerably small measurement samples can be used in the modeling process.

Erythropoietin promotes retinal angiogenesis in a mouse model.

This study aimed to investigate the effect and potential mechanisms of exogenous administration of recombinant human erythropoietin (rhEPO) on retinal angiogenesis in a mouse model of oxygen-induced retinopathy (OIR). Postnatal day 7 (P7) mice (n=132) were randomly assigned to one of six groups: Control group (n=22), OIR group (n=22), OIR + vehicle control group (n=22), OIR + rhEPO 10 IU group (n=22), OIR + rhEPO 50 IU group (n=22), and OIR + rhEPO 100 IU group (n=22). OIR was induced by exposing mice to 75±2% O2 for five days, followed by exposure to room air for a further five days. Animals in groups 3-6 (the OIR + vehicle control group and OIR + rhEPO 10 IU, 50 IU, and 100 IU groups) received an intraperitoneal injection of saline, or rhEPO 10 IU, 50 IU and 100 IU, respectively, which were administered daily from P7-P12. Immunofluorescent and hematoxylin-eosin staining were used to detect retinal neovascularization (RNV) in retinal whole mounts. Quantitative polymerase chain reaction and western blot analysis were used to detect the expression levels of vascular endothelial growth factor (VEGF), endothelial nitric oxide synthase (eNOS), and neuronal nitric oxide synthase (nNOS). RNV occurred in the OIR groups and was accompanied by dilated, twisted and occluded blood vessels. rhEPO treatment resulted in an increase in the number of newly formed and severely dilated vessels. rhEPO increased RNV in a dose-dependent manner, which was accompanied by an increase in the messenger RNA and protein expression of VEGF, eNOS and nNOS. Thus, exogenous use of rhEPO promotes the RNV in a mouse model of OIR and is accompanied by increased expression levels of VEGF, eNOS and nNOS.

Poster Presentations

Administration of recombinant human erythropoietin (rHuEpo) improves performance and hence is subject to abuse by athletes. The detection of rHuEpo doping remains a major challenge at present. The aim of the current study was to investigate whether circulating microRNA (miRNA) can be used for detecting r-HuEpo doping. Twenty trained males received rHuEpo injections of 50 IU∙kg -1 body mass every two days for 4 weeks. Blood was obtained 2 weeks before, during and 4 weeks after administration. For this pilot analysis, plasma miRNA expression was assessed at selected time points using the Affymetrix GeneChip 3.0 and the miScript 384 HC PCR Array (Qiagen). For the Affymetrix microarray data, GC content adjustment was carried out prior to background correction and quantile normalization. For the Qiagen PCR Array data, raw CT values exported from the RQ manager were firstly normalized to the spike-in control (i.e. cel-miR-39-3p), then to the mean of commonly expressed miRNAs with a CT < 35 prior to formal analysis. 1733 human miRNAs are present on the GeneChip 3.0. Eighty-two of the 1733 miRNAs were commonly expressed across all samples and 773 miRNAs were found in at least one sample. Six miRNAs showed significant fold changes following r-HuEpo, with 5 miRNAs up-regulated (fold change ranging from 2.03-3.07, P < 0.05) and 1 miRNA down-regulated (fold change: 2.56, P = 0.042). Out of 372 human miRNAs present (Qiagen), 205 human miRNAs were detected in all samples and 164 miRNAs were found in at least one sample. Two miRNAs were differentially expressed (fold changes: 3.4 - 3.84, P ≤ 0.03) following r-HuEpo. For significantly expressed miRNAs identified by both methods, 6 common miRNAs were found to be either predicted or validated miRNAs potentially involved in Epo signalling pathways. Analysis of identified miRNAs in the remaining samples will be required in order to determine whether circulating miRNAs can be effectively used for r-HuEpo doping detection. miRNAs isolated from red blood cells before and after r-HuEpo are also currently being analysed using next generation sequencing.

The Efficacy of Intravenous versus Subcutaneous Recombinant Erythropoietin in Obese African-African Patients in a Southeast U.S. Dialysis Cohort

Aims: To correct renal anemia, subcutaneous (SC) route of recombinant human erythropoietin (rhuEPO) administration has been associated with increased efficacy and decreased dose requirements, when compared with intravenous (IV) route. The effect of obesity as a potential modifier during rhuEPO administration has not been well explored. Study Design: Single-center, Longitudinal Cohort Study. Place and Duration of Study: University of Mississippi Medical Center Outpatient Dialysis Unit, between February and November of 2009. Research Article British Journal of Medicine & Medical Research, 4(1): 184-193, 2014 185 Methodology: We performed IV to SC rhuEPO conversion for 86 in-center dialysis patients and, following a six-month equilibration period, we monitored outcomes over a period of three months. We obtained baseline demographic parameters, calculated Body Mass Index (BMI) and monitored iron saturation, ferritin, hemoglobin (Hgb) along with rhuEPO requirements. Patients were divided into 3 categories based on BMI [ 35 (n= 21) kg/m]. Results are reported either as percents, means with SD or median with 25-75% interquartile range, as appropriate. Results: The cohort was all African-American, 48.8% male, aged 54.7 (13.3) years and BMI calculated at 29.9 (7.4) kg/m. Baseline iron saturation was 24 (10.6)%, ferritin measured 641 (277) ng/mL. Hgb remained unchanged during the observation period: 11.1 (1.3) vs. 11.2 (1.3) gm/dL. Initial rhuEPO weekly dose for the entire cohort was 19,729 (17,448) Units/week (U/week); final dose 17,482 (14,860) U/week, with close correlation between initial and final doses (r: 0.653, P<0.0001). Weekly rhuEPO dose remained virtually unchanged in BMI categories 1 and 2 [13,927 (10,938) vs. 13,297 (10,247) U/week; 20,684 (15,788) vs. 20,997 (17.917)] (P=NS for both) but decreased in the category 3: 25,459 (24,403) vs. 16,444 (12,749) (P=0.081). However, BMI had no independent effect in linear regression modeling with multiple covariates (age, BMI, iron saturation, ferritin) included. Conclusion: Obesity may affect relative efficacy of rhuEPO conversion; additional studies may be needed.

Does bleeding induce micronuclei via erythropoietin in Han-Wistar rats?

The effects of both intravenous administration of recombinant human erythropoietin (rhEPO) and blood withdrawal, either individually or in combination, on the frequency of micronucleated immature erythrocytes (MIE) in the bone marrow were investigated in 10-week old male Han Wistar rats. Clear increases in MIE frequency and evidence of erythroid hyperplasia were seen in the bone marrow after 1350 IU kg−1 rhEPO but only marginal increases in MIE were seen at the lower dose of 135 IU kg−1. At the lower dose, blood levels of rhEPO were found to be approximately 120 pg mL−1 within one hour of dosing. Withdrawal of 5 blood samples of 0.45 mL on each occasion during 24 hours, the maximum allowed by the Home Office licence, did not increase MIE frequency but did result in levels of endogenous EPO of 15–19 pg mL−1 24 to 72 hours after the start of treatment, with some evidence of increased erythropoiesis. Although the increased plasma amounts of EPO, resulting from blood withdrawal, were lower than the blood levels achieved by the administration of exogenous rhEPO that were required to give increases in micronucleus frequency, the possibility that blood withdrawal might influence the MIE response to genotoxins cannot be totally excluded without further investigation.

Erythropoietin in cardiac disease

Discovered as the primary regulator of erythropoiesis, erythropoietin (EPO) is involved in a broad variety of processes that play a major role in cardiovascular diseases. In particular, the antiapoptotic and pro-angiogenic properties of EPO have prompted a growing interest in the use of EPO for the treatment of myocardial infarction and heart failure. In a variety of myocardial ischemic injury animal models, EPO administration has been shown to acutely reduce infarct size, thereby preserving ventricular function. In addition, cardiac long-term effects of EPO, such as prevention of ventricular remodeling and heart failure, have been described. In recent years, several trials have tested the effects of recombinant human erythropoietin (rhEPO) administration in patients with myocardial infarction and chronic heart failure, in the attempt to translate the cardioprotection found in experimental models to human patients. In view of the generally controversial findings, in this updated review we provide an overview of the results of the most recent trials that investigated the role of erythropoiesis-stimulating agents (ESAs), including rhEPO and its analogue darbepoetin, in the treatment of acute myocardial infarction and heart failure. The problems related to safety and tolerability of ESA therapy are also discussed. Our analysis of the available literature demonstrates that the results of clinical studies in patients with cardiac disease are not uniform and the conclusions are contradictory. Further larger prospective studies are required to test clinical efficacy and safety of EPO.

Iron depletion induced by bloodletting and followed by rhEPO administration as a therapeutic strategy in progressive multiple sclerosis: A pilot, open-label study with neurophysiological measurements

To evaluate the concept that iron depletion (ID) induced by bloodletting and followed by recombinant human erythropoietin (rhEPO) administration could be a therapeutic strategy in progressive multiple sclerosis (PMS) and that it could be assessed by neurophysiological measurements. In four patients with PMS, bloodletting was performed until ID was induced, and then rhEPO was administered (300 UI/kg/week). The changes induced by the treatment were assessed by clinical scores, biological tests, and neurophysiological study of cortical excitability using transcranial magnetic stimulation techniques. The treatment was well tolerated except for muscle cramps and one popliteal vein thrombosis in a patient confined to chair. ID was obtained within 28 weeks and was associated with endogenous production of EPO. No bloodletting was further required during a six-month period after introduction of rhEPO. At the end of the follow-up (up to one year), fatigue and walking capacities tended to improve in two patients. Neurophysiological changes were characterized by an increased cortical excitability, including a decrease of motor thresholds and an enhancement of intracortical facilitation and cerebellothalamocortical inhibition. The combined ID-rhEPO therapy could authorize a prolonged administration of rhEPO in PMS patients, able to modify cortical excitability of the glutamatergic and gabaergic circuits. These preliminary data are encouraging to design a larger, controlled therapeutical trial to assess the value of such a strategy to improve functional symptoms in PMS patients, and maybe to prevent axonal degeneration. Neurophysiological measurements based on cortical excitability studies could provide sensitive parameters to evaluate treatment-induced changes in this context.