losartan-potassium and Reperfusion-Injury

Stroke is a debilitating disease and has the ability to culminate in devastating clinical outcomes. Ischemic stroke followed by reperfusion entrains cerebral ischemia/reperfusion (I/R) injury, which is a complex pathological process and is associated with serious clinical manifestations. Therefore, the development of a robust and effective poststroke therapy is crucial. Granulocyte colony-stimulating factor (GCSF) and erythropoietin (EPO), originally discovered as hematopoietic growth factors, are versatile and have transcended beyond their traditional role of orchestrating the proliferation, differentiation, and survival of hematopoietic progenitors to one that fosters brain protection/neuroregeneration. The clinical indication regarding GCSF and EPO as an auspicious therapeutic strategy is conferred in a plethora of illnesses, including anemia and neutropenia. EPO and GCSF alleviate cerebral I/R injury through a multitude of mechanisms, involving antiapoptotic, anti-inflammatory, antioxidant, neurogenic, and angiogenic effects. Despite bolstering evidence from preclinical studies, the multiple brain protective modalities of GCSF and EPO failed to translate in clinical trials and thereby raises several questions. The present review comprehensively compiles and discusses key findings from in vitro, in vivo, and clinical data pertaining to the administration of EPO, GCSF, and other drugs, which alter levels of colony-stimulating factor (CSF) in the brain following cerebral I/R injury, and elaborates on the contributing factors, which led to the lost in translation of CSFs from bench to bedside. Any controversial findings are discussed to enable a clear overview of the role of EPO and GCSF as robust and effective candidates for poststroke therapy.

Topics: Animals; Colony-Stimulating Factors; Erythropoietin; Humans; Ischemic Stroke; Reperfusion Injury

The renoprotection of erythropoietin (EPO) and its derivatives such as helix B surface peptide (HBSP) have attracted a great deal of attention from scientists and clinicians alike. The evolutional achievement in the dissociation of tissue protection and erythropoiesis is obtained through HBSP characterisation and synthesis. We performed a series of studies using EPO, as well as HBSP, in a variety of biological models subjected to transplant-related renal injuries such as ischemia reperfusion injury (IRI) and/or immunosuppressant nephrotoxicity. In this short review, we would like to address the effects of EPO in different formats, and its underlying mechanisms with focuses on apoptosis and inflammation in in vitro, ex vivo and in vivo renal injury models, and to further explore potential applications and challenges in humans.

Topics: Acute Kidney Injury; Animals; Anti-Inflammatory Agents; Apoptosis; Cytokine Receptor Common beta Subunit; Disease Models, Animal; Erythropoietin; Gene Expression Regulation; Humans; Kidney; Oxidative Stress; Peptide Fragments; Protective Agents; Receptors, Erythropoietin; Reperfusion Injury

Erythropoietin (EPO), recognized early as a tissue protective agent, can trigger antiinflammatory and anti-apoptotic processes to delimit injury and promote repair by binding tissueprotective receptor (TPR). However, only at a high dosage can EPO exert tissue protective effect, which may elicit severe side-effects at the meantime. Helix B surface peptide (HBSP), a 11-amnio acid sequence derived from the non-erythropoietic helix B of EPO, not only shows higher affinity to TPR but also plays a more specific and powerful role in tissue protection without erythropoietic side-effects. While it has obvious merits, the 2-min plasma half-life of HBSP restricts its application in vivo. Therefore, based on the amino acid sequence of HBSP, we originally designed and synthesized thioethercyclized helix B peptide (CHBP) for an increased resistance to proteolytic degradation as well as an improved tissue protective potency, implying a brighter prospective for translational application. In this review, we will mainly discuss the development from EPO to CHBP, the merits and limitation of CHBP and the probable mechanism mediating tissue protection.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; Cyclization; Cytokine Receptor Common beta Subunit; Disease Models, Animal; Erythropoietin; Gene Expression Regulation; Half-Life; Humans; Kidney; Oxidative Stress; Peptide Fragments; Peptides, Cyclic; Protective Agents; Receptors, Erythropoietin; Reperfusion Injury

Renal ischemia reperfusion injury (IRI) contributes to the development of acute kidney injury (AKI). Several processes are involved in the development of renal IRI with the generation of reactive oxygen species, inflammation and apoptosis. MicroRNAs (miRNAs) are endogenous, small and noncoding RNAs that repress gene expression of target mRNA in animals post-transcriptionally. miRNA-mediated gene repression is a major modulatory mechanism to regulate fundamental cellular processes such as the cell cycle, proliferation, growth, and apoptosis, which in turn have pivotal influences on pathophysiological outcomes. Recent studies have revealed the pathogenic roles played by miRNAs in many renal diseases, such as IRI, AKI and renal carcinoma. In addition, the majority of miRNAs identified appear to be differentially expressed, probably to quell the injury response by modulating inflammation, apoptosis and proliferation and may point us toward new pathways that can be targeted to regulate or prevent renal IRI. They may represent novel diagnostic biomarkers of renal IR injury.

Topics: Acute Kidney Injury; Animals; Apoptosis; Erythropoietin; Humans; Melatonin; Mice; MicroRNAs; Rats; Reactive Oxygen Species; Reperfusion Injury

The protective, nonerythropoietic effects of erythropoietin (EPO) have become evident in preclinical models in renal ischaemia/reperfusion injury and kidney transplantation. However, four recently published clinical trials using high-dose EPO treatment following renal transplantation did not reveal any protective effect for short-term renal function and even reported an increased risk of thrombosis. This review focusses on the current status of protective pathways mediated by EPO, the safety concerns using high EPO dosage and discusses the discrepancies between pre-clinical and clinical studies. The protective effects are mediated by binding of EPO to a heteromeric receptor complex consisting of two β-common receptors and two EPO receptors. An important role for the activation of endothelial nitric oxide synthase is proposed. EPO-mediated cytoprotection still has enormous potential. However, only nonerythropoietic EPO derivatives may induce protection without increasing the risk of cardiovascular events. In preclinical models, nonerythropoietic EPO derivatives, such as carbamoylated EPO and ARA290, have been tested. These EPO derivatives improve renal function and do not affect erythropoiesis. Therefore, nonerythropoietic EPO derivatives may be able to render EPO-mediated cytoprotection useful and beneficial for clinical transplantation.

Topics: Animals; Cardiovascular Diseases; Cytoprotection; Erythropoietin; Hematinics; Humans; Kidney; Kidney Transplantation; Nitric Oxide Synthase Type III; Oligopeptides; Receptors, Erythropoietin; Recombinant Proteins; Reperfusion Injury; Risk Factors; Translational Research, Biomedical

Testicular torsion is a urological emergency most commonly seen in adolescence, involving a decrease in blood flow in the testis resulting from torsion of the spermatic cord that can result in gonad injury or even loss if not treated in time. Testicular ischaemia-reperfusion injury represents the principle pathophysiology of testicular torsion, with ischaemia caused by twisting of the spermatic cord, and reperfusion on its subsequent release. Many cellular and molecular mechanisms are involved in ischaemia-reperfusion injury following testicular torsion. Studies have investigated the use of pharmacological agents as supportive therapy to surgical repair in order to prevent the adverse effects of testicular torsion. Numerous substances have been proposed as important in the prevention of post-ischaemia-reperfusion testicular injury. A range of chemicals and drugs has been successfully tested in animal models for the purpose of mitigating the dangerous effects of ischaemia-reperfusion in testis torsion.

Topics: Adjuvants, Immunologic; Anesthetics; Animals; Anti-Inflammatory Agents, Non-Steroidal; Antioxidants; Dehydroepiandrosterone; Drug Therapy, Combination; Erythropoietin; Humans; Hyperbaric Oxygenation; Male; Phosphodiesterase Inhibitors; Reperfusion Injury; Spermatic Cord Torsion; Treatment Outcome; Urologic Surgical Procedures, Male; Vasodilator Agents

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.

Topics: Animals; Epoetin Alfa; Erythropoietin; Humans; Neovascularization, Physiologic; Nitric Oxide Synthase Type II; Recombinant Proteins; Regeneration; Reperfusion Injury; Skin; Skin Physiological Phenomena; Surgical Flaps; Vascular Endothelial Growth Factor A; Wound Healing

Sudden cardiac arrest is a leading cause of death worldwide with survival rates still remaining suboptimal. Unfortunately, most cardiac arrest patients, who achieve return of spontaneous circulation (ROSC), develop a multi-faceted post-cardiac arrest syndrome, including post-cardiac arrest brain injury, myocardial dysfunction, and systemic ischemia/reperfusion response. Erythropoietin (EPO), the principal hematopoietic hormone regulating erythropoiesis, exhibits diverse cellular effects in nonhematopoietic tissues. Due to its anti-apoptotic, anti-inflammatory, and anti-oxidant properties, as well as its angiogenic action, EPO plays a role in neuroprotection and cardioprotection. In this regard, EPO represents a promising agent in the cardiac arrest setting, based on a therapeutic strategy that focuses on the post-resuscitation phase. This review aims to provide a comprehensive account of EPO's role in the treatment of each individual component of post-cardiac arrest syndrome.

Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Apoptosis; Brain Diseases; Cardiomyopathies; Cardiotonic Agents; Erythropoietin; Heart Arrest; Humans; Reperfusion Injury; Syndrome

Erythropoietin (EPO) is a well-known therapeutic protein employed widely in the treatment of anemia. Over the past decade, abundant evidence has shown that in addition to its systemic role in the regulation of plasma pO(2) by modulating erythrocyte numbers, EPO is also a cytoprotective molecule made locally in response to injury or metabolic stress. Many studies have shown beneficial effects of EPO administration in reducing damage caused by ischemia-reperfusion, trauma, cytotoxicity, infection and inflammation in a variety of organs and tissues. Notably, the receptor mediating the nonerythropoietic effects of EPO differs from the one responsible for hematopoiesis. The tissue-protective receptor exhibits a lower affinity for EPO and is a heteromer consisting of EPO receptor monomers in association with the common receptor that is also employed by granulocyte macrophage colony-stimulating factor, interleukin 3, and interleukin 5. This heteromeric receptor is expressed immediately following injury, whereas EPO production is delayed. Thus, early administration of EPO can dramatically reduce the deleterious components of the local inflammatory cascade. However, a high dose of EPO is required and this also stimulates the bone marrow to produce highly reactive platelets and activates the vascular endothelium into a prothrombotic state. To circumvent these undesirable effects, the EPO molecule has been successfully altered to selectively eliminate erythropoietic and prothrombotic potencies, while preserving tissue-protective activities. Very recently, small peptide mimetics have been developed that recapitulate the tissue-protective activities of EPO. Nonerythropoietic tissue-protective molecules hold high promise in a wide variety of acute and chronic diseases.

Topics: Amino Acid Sequence; Animals; Cytokine Receptor Common beta Subunit; Cytokines; Drug Design; Erythropoietin; Feedback, Physiological; Hematinics; Hematopoiesis; Humans; Inflammation; Inflammation Mediators; Janus Kinase 2; Models, Molecular; Molecular Sequence Data; Protein Conformation; Protein Multimerization; Receptors, Erythropoietin; Recombinant Proteins; Reperfusion Injury; STAT Transcription Factors; Stress, Physiological; Wounds and Injuries

Analysis of published data indicates that erythropoietin plays an important role in regulation of brain tolerance to impact of ischemia-reperfusion. This cytokine is involved in ischemic preconditioning of the brain and can mimic the phenomenon of preconditioning and postconditioning. However, it is unclear whether endogenous erythropoietin takes part in postconditioning of the brain. It is established that erythropoietin inhibits apoptosis of neurons after ischemia-reperfusion. It was found that erythropoietin could stimulate regeneration of the brain after ischemia. Signaling mechanism of neuroprotective action of erythropoietin is well studied but there are very little data on the mechanism of erythropoietin-induced neurogenesis and neoangiogenesis. It is unknown whether erythropoietin can exhibit infarct-limiting effect in humans and stimulate neurogenesis and neoangiogenesis in patients after insult.

Topics: Animals; Apoptosis; Brain; Brain Infarction; Erythropoietin; Ischemic Preconditioning; Male; Neovascularization, Physiologic; Neurons; Regeneration; Reperfusion Injury; Stroke

Erythropoietin is a hypoxia-induced cytokine that stimulates erythropoiesis through the promotion of erythroid precursor cell proliferation and differentiation. Recent evidence supports that erythropoietin has a broad spectrum of tissue protecting actions affecting other systems than hemopoietic. Lately, research has focused on the nonhemopoietic effects of erythropoietin against tissue ischemia due to the unexpected observations of erythropoietin receptor expression by various cells, such as endothelial cells, neuronal cells, cardiac myocytes, and vascular smooth muscle cells. It has been shown that erythropoietin exerts its cardioprotective action during cardiac ischemic injury through reducing the infract size and enhancing new vessel formation over a longer time frame. Erythropoietin plays a crucial role in neuroprotection in many types of ischemic injury in the central and the peripheral nervous system. It is also strongly believed that erythropoietin exhibits a critical role in many other disorders that are pathogenetically related to acute tissue ischemia. This article reviews the proposed implications of erythropoietin in tissue ischemia and discusses the possible mechanisms for this action along with its potential therapeutic applications.

Topics: Animals; Apoptosis; Cardiovascular System; Cell Survival; Endothelial Cells; Erythropoietin; Humans; Hypoxia; Ischemia; Models, Biological; Nervous System; Neurons; Oxygen; Reperfusion Injury

This study reviews the current understanding of ischemic preconditioning (IP) in experimental and clinical setting, and the mechanisms that mediate the complex processes involved as a tool to protect against ischemia and reperfusion (I/R) injury, but is not intended as a complete literature review of preconditioning. IP has been mainly elucidated in cardiac ischemia. Recent reports confirm the efficacy of pre- and postconditioning in cardiac surgery and percutaneous coronary interventions in humans. IP utilizes endogenous as well as distant mechanisms in skeletal muscle, liver, lung, kidney, intestine and brain in animal models to convey varying degrees of protection from I/R injury. Specifically, preconditioned tissues exhibit altered energy metabolism, better electrolyte homeostasis and genetic reorganization, as well as less oxygen-free radicals and activated neutrophils release, reduced apoptosis and better microcirculatory perfusion. To date, there are few human studies, but recent trials suggest that human liver, lung and skeletal muscle acquire protection after IP. Present data address the potential therapeutic application of IP in the prevention of I/R damage specially aimed at clinical transplantation. IP is ubiquitous but more research is required to fully translate these findings to the clinical arena.

Topics: Animals; Erythropoietin; Humans; Ischemic Preconditioning; Kidney Transplantation; Liver Transplantation; Organ Transplantation; Reperfusion Injury

Renal ischemia-reperfusion (I-R) contributes to the development of ischemic acute renal failure (ARF). Multi-factorial processes are involved in the development and progression of renal I-R injury with the generation of reactive oxygen species, nitric oxide and peroxynitrite, and the decline of antioxidant protection playing major roles, leading to dysfunction, injury, and death of the cells of the kidney. Renal inflammation, involving cytokine/adhesion molecule cascades with recruitment, activation, and diapedesis of circulating leukocytes is also implicated. Clinically, renal I-R occurs in a variety of medical and surgical settings and is responsible for the development of acute tubular necrosis (a characteristic feature of ischemic ARF), e.g., in renal transplantation where I-R of the kidney directly influences graft and patient survival. The cellular mechanisms involved in the development of renal I-R injury have been targeted by several pharmacological interventions. However, although showing promise in experimental models of renal I-R injury and ischemic ARF, they have not proved successful in the clinical setting (e.g., atrial natriuretic peptide, low-dose dopamine). This review highlights recent pharmacological developments, which have shown particular promise against experimental renal I-R injury and ischemic ARF, including novel antioxidants and antioxidant enzyme mimetics, nitric oxide and nitric oxide synthase inhibitors, erythropoietin, peroxisome-proliferator-activated receptor agonists, inhibitors of poly(ADP-ribose) polymerase, carbon monoxide-releasing molecules, statins, and adenosine. Novel approaches such as recent research involving combination therapies and the potential of non-pharmacological strategies are also considered.

Topics: Acute Kidney Injury; Animals; Antioxidants; Erythropoietin; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Inflammation; Kidney; Kidney Diseases; Nitric Oxide; Nitric Oxide Synthase; Peroxisome Proliferator-Activated Receptors; Poly(ADP-ribose) Polymerase Inhibitors; Purinergic P1 Receptor Agonists; Reactive Nitrogen Species; Reactive Oxygen Species; Receptors, Lysosphingolipid; Recombinant Proteins; Reperfusion Injury

A greatly expanded understanding of the biology of endogenous erythropoietin (EPO) has emerged since the early 1990s. Originally viewed as the renal hormone dedicated to erythrocyte production, it is now clear that EPO is produced locally by many other tissues in response to physical or metabolic stress. In its autocrine-paracrine roles, EPO mediates preconditioning (ischemic tolerance) and specifically limits the destructive potential of tumor necrosis factor alpha and other proinflammatory cytokines in the brain, heart, kidney, and other tissues. As local production of EPO is generally suppressed following injury, administration of exogenous EPO has been a successful therapeutic approach in preclinical and clinical studies, for example, following ischemia-reperfusion and toxin-induced renal injuries, and in human stroke. The therapeutic time window of tissue protection by EPO is typically very wide in experimental models, showing effectiveness when administered before, during, or after an insult and raising optimism for a high clinical potential. Although there is progress in understanding the signaling pathways responsible for EPO's tissue-protective actions that are similar to, but not as redundant as, those employed for erythrocyte maturation, much work remains to be carried out. Experimental observations also suggest the existence of EPO receptor (EPOR) isoforms mediating EPO's diverse biological activities and have identified a tissue-protective receptor complex consisting of the EPOR and the beta common receptor (CD131) subunit that is also employed by granulocyte-macrophage colony-stimulating factor, interleukin-3 and interleukin-5. Successfully engineered analogues of EPO that selectively activate tissue protection without stimulating hematopoiesis confirm the concept of a tissue-protective receptor and have significant potential utility in the investigational and therapeutic arenas.

Topics: Animals; Endothelium, Vascular; Erythropoietin; Humans; Kidney; Kidney Diseases; Reperfusion Injury

The haematopoietic factor erythropoietin (EPO) has recently been recognized to play a physiological role in the brain and other tissues. The EPO receptor is present in the glomerulus, mesangial and tubular epithelial cells in the kidney. We have reviewed the experimental use of EPO in animal models of acute renal failure. EPO attenuates the dysfunction and histological changes associated with ischaemia-reperfusion injury, with a reduction in apoptotic cell death. EPO has also shown benefit in animal models of systemic shock and cisplatin-induced nephrotoxicity. In vitro studies have shown that EPO has direct effects on proliferation and cell death in proximal tubular epithelial cells. There is increasingly strong experimental evidence that EPO may be of therapeutic use in acute renal failure, and clinical trials should be undertaken to determine its clinical applications in this field.

Topics: Acute Kidney Injury; Animals; Cytoprotection; Endothelium, Vascular; Erythropoietin; Kidney; Reperfusion Injury

The hemopoietic growth factor erythropoietin (EPO) has been recognized to be a multifunctional cytokine that plays a key role in ischemic preconditioning in the brain and heart. The EPO receptor is expressed widely in the kidney, and we review the important findings from the use of EPO in experimental models of acute renal failure that show that EPO reduces tubular cell death and hence the dysfunction induced by ischemia reperfusion injury, and we explore how these observations may be translated into the clinical arena.

Topics: Acute Kidney Injury; Animals; Apoptosis; Brain Ischemia; Erythropoietin; Humans; Kidney; Kidney Tubules, Proximal; Recombinant Proteins; Reperfusion Injury

Cardiac surgery and cardiopulmonary bypass (CPB) induce ischemia-reperfusion and subsequent cellular injury with inflammatory reaction. Clinical and experimental studies suggest that recombinant human erythropoietin (EPO) independently of its erythropoietic effect may be used as a cytoprotective agent against ischemic injury. We tested the hypothesis that one large dose of EPO administered shortly before CPB prevents the elevation of cardiac and cerebral ischemic blood markers as well as the systemic inflammatory response induced by cardiac surgery with CBP through this randomized double-blind placebo-controlled pilot trial. Fifty patients scheduled for coronary artery bypass graft (CABG) surgery with CPB were randomly allocated to EPO or control groups. EPO (800 IU/kg intravenously) or placebo (saline) was administered before CPB. The primary end point was to study the effect of EPO administration on several blood markers of myocardial and cerebral ischemia in relation to CABG with CPB. In both groups, surgery increased plasma concentrations of cardiac (troponin T, NT-proBNP, and creatine kinase MB) and cerebral (S100β protein) markers ischemic as well as the pro-inflammatory marker interleukin-6. Compared with the placebo, EPO administration before CPB did not prevent an increase of all these markers following CPB. In conclusion, one large dose of EPO, given shortly before CPB, did not protect against cardiac and cerebral ischemia and inflammatory response occurring during CABG surgery with CPB. Although the long-term clinical implications remain unknown, the findings do not support use of EPO at this dose as a cytoprotective agent in patients undergoing cardiac surgery.

Topics: Aged; Biomarkers; Brain Ischemia; Cardiopulmonary Bypass; Cytokines; Cytoprotection; Data Interpretation, Statistical; Dose-Response Relationship, Drug; Double-Blind Method; Electrocardiography; Erythropoietin; Female; Humans; Male; Middle Aged; Myocardial Ischemia; Myocardial Reperfusion Injury; Nerve Growth Factors; Pilot Projects; Recombinant Proteins; Reperfusion Injury; S100 Calcium Binding Protein beta Subunit; S100 Proteins; Treatment Outcome; Troponin T

To determine the effect of recombinant erythropoietin on serum oxidants and the viability of ischemic ovaries after detorsion.. A non-randomized single-blind clinical trial was conducted from December 2009 to January 2011 in a University Teaching Hospital affiliated with the School of Medicine, Tabriz University of Medical Sciences. Surgery was carried out on 40 patients, aged 18-35 years, with signs and symptoms of ovarian torsion. The patients were divided into two equal groups: group 1 received recombinant erythropoietin 150 IU/kg subcutaneously during the operation and 72 h after detorsion, and group 2 received no medication. Blood samples were taken before and 72 h after detorsion to determine the plasma levels of malondialdehyde, xanthine oxidase, glutathione, superoxide dismutase, nitric oxide, and total antioxidants. In both groups, the arterial and venous blood supply of the ovary and arterial blood flow resistance were evaluated before surgery and 72 h after detorsion of the ovary. The main outcome measures were improving ovarian blood flow and reducing oxidative damage. SPSS 17.0 was used for statistical analyses.. The levels of malondialdehyde, glutathione, superoxide dismutase, nitric oxide, and total antioxidants 72 h after detorsion were significantly different between the interventional and non-interventional groups (p<0.001). There was no significant difference in the levels of xanthine oxidase (p=0.13). The difference between groups in the blood flow of the ovary 72 h after surgery was not statistically significant (p=0.61).. Recombinant erythropoietin was effective in reducing the oxidative damage of ovarian torsion.

Topics: Adolescent; Adult; Erythropoietin; Female; Humans; Ovarian Diseases; Oxidative Stress; Postoperative Complications; Recombinant Proteins; Reperfusion Injury; Torsion Abnormality; Young Adult

Delayed graft function (DGF) is associated with adverse long-term outcomes after deceased-donor kidney (DDK) transplantation. Ischemia-reperfusion injury plays a crucial role in the development of DGF. On the basis of promising animal data, this study evaluated any potential benefits of erythropoietin-alfa (EPO-α) given intra-arterially at the time of reperfusion of renal allograft on the degree of allograft function, as well as tubular cell injury measured by urinary biomarkers in the early post-transplant period.. A prospective, randomized, double-blind, placebo-controlled clinical trial was conducted to evaluate the influence of EPO-α administered intraoperatively on the outcomes of DDK transplantations performed at the study center between March 2007 and July 2009.. Seventy-two patients were randomly assigned to EPO-α (n=36) or placebo (n=36). The incidences of DGF, slow graft function, and immediate graft function did not significantly differ between the treatment and control groups (41.7% versus 47.2%, 25.0% versus 36.1%, and 33.3% versus 16.7%, respectively; P=0.24). The groups had similar levels of urinary biomarkers, including neutrophil gelatinase-associated lipocalin and IL-18 at multiple times points soon after transplantation; urinary output during the first 3 postoperative days; 1-month renal function; and BP readings, hemoglobin, and adverse effects during the first month.. This study did not show any clinically demonstrable beneficial effects of high-dose EPO-α given intra-arterially during the early reperfusion phase in DDK transplant recipients in terms of reducing the incidence of DGF or improving short-term allograft function.

Topics: Acute-Phase Proteins; Adult; Aged; Analysis of Variance; Biomarkers; Blood Pressure; Chi-Square Distribution; Creatinine; Delayed Graft Function; Double-Blind Method; Drug Administration Schedule; Epoetin Alfa; Erythropoietin; Female; Hemoglobins; Humans; Incidence; Injections, Intra-Arterial; Interleukin-18; Kidney Transplantation; Lipocalin-2; Lipocalins; Male; Middle Aged; Pennsylvania; Prospective Studies; Proto-Oncogene Proteins; Recombinant Proteins; Reperfusion Injury; Time Factors; Treatment Outcome; Young Adult

The development of novel pharmaceutical interventions to improve the clinical outcomes of patients with acute ST-segment elevation myocardial infarction (STEMI) is an unmet medical need worldwide. In animal models, a single intravenous administration of erythropoietin (EPO) during reperfusion improves left ventricular (LV) function in the chronic stage. However, the results of recent proof-of-concept trials using high-dose EPO in patients with STEMI are inconsistent. In our pilot study, low-dose EPO after successful percutaneous coronary intervention (PCI) improved the LV ejection fraction (EF) and did not trigger severe adverse clinical events in patients with STEMI. One possible reason for this discrepancy is the dose of EPO used.. We have started a double-blind, placebo-controlled, randomized, multicenter clinical trial (EPO-AMI-II) to clarify the safety and efficacy of low-dose EPO in patients with STEMI. STEMI patients who have a low LVEF (<50 %) will be randomly assigned to intravenous administration of placebo or EPO (6,000 or 12,000 IU) within 6 h after successful PCI. The primary endpoint is the difference in LVEF between the acute and chronic phases (6 months), as measured by single-photon emission computed tomography. The patient number needed for EPO-AMI-II is 600. The study will stop when superior efficacy or futility is detected by an interim analysis. This study has been approved by the Evaluation System of Investigational Medical Care.. EPO-AMI-II study will clarify the safety and efficacy of low-dose EPO in STEMI patients with LV dysfunction in a double-blind, placebo-controlled, multicenter study. (247 words).

Topics: Adult; Aged; Double-Blind Method; Erythropoietin; Humans; Middle Aged; Myocardial Infarction; Percutaneous Coronary Intervention; Reperfusion Injury; Ventricular Dysfunction, Left; Young Adult

To investigate the protective effect of erythropoietin (Epo) against ischemia-reperfusion injury (IR/I) following the Pringle maneuver (PM), in comparison with conventional steroid administration in a prospective randomized trial.. Patients were randomized by age, sex, diagnosis, and surgical method, and assigned to three groups: (1) A steroid group (STRD, n = 9) who received 100 mg of hydrocortisone before PM, and on postoperative days 1, 2 and 3, followed by tapering until postoperative day 7; (2) An EPO1 group (n = 10) who received 30,000 U of Epo before the PM and at the end of surgery; and (3) An EPO2 group (n = 8) who received 60,000 U of Epo before the PM. Hemoglobin (Hb), hematocrit (Ht), aspartate aminotransferase (AST), alanine transaminase (ALT), lactate dehydrogenase (LDH), lactate, interleukin-6 (IL-6), and tumor necrosis factor (TNF)-α were measured before and just after (Day 0) surgery, and on postoperative days 1, 3, 7 and 14.. There were no increases in Hb and Ht in the EPO1 and EPO2 groups. AST was significantly lower in EPO1 than in STRD on Day 0 (P = 0.041), and lower in EPO1 than in STRD and EPO2 on Day 1 (P = 0.018). ALT was significantly lower in EPO1 than in STRD and EPO2 on Day 0 (P = 0.020) and Day 1 (P = 0.004). There were no significant inter-group differences in the levels of LDH and lactate. IL-6 was significantly lower in EPO1 than in STRD and EPO2 on Day 0 (P = 0.0036) and Day 1 (P = 0.0451). TNF-α was significantly lower in EPO1 than in STRD and EPO2 on Day 0 (P = 0.0006) and Day 1 (P < 0.0001). Furthermore, hospitalization was significantly shorter in EPO1 and EPO2 than in STRD.. Epo has greater potential than steroids to ameliorate IR/I after the PM. Epo at a dose of 30,000 U, administered before PM and just after surgery, yields better results.

Topics: Aged; Animals; Digestive System Surgical Procedures; Erythropoietin; Female; Hematocrit; Hemoglobins; Humans; L-Lactate Dehydrogenase; Lactic Acid; Length of Stay; Liver Diseases; Male; Middle Aged; Prospective Studies; Reperfusion Injury; Steroids; Tumor Necrosis Factor-alpha

Erythropoietin (EPO) exerts a tissue-protective activity in several non-haematopoietic tissues such as heart, brain, spinal cord and muscle. We evaluated the relationship between pre-operative endogenous EPO blood levels and myocardial damage in patients undergoing cardiopulmonary bypass (CPB). Furthermore, we investigated whether pre-operative administration of a single bolus of 40,000 IU epoetin alpha (EPOalpha) would reduce troponin I or creatine kinase isoenzyme (CK-MB) after on-pump coronary artery bypass graft (CABG) surgery. Sixty-seven patients (45 CABG, 22 valvular surgery) were enrolled. EPO was measured in the pre-surgical period and correlated to post-surgical troponin I and CK-MB peaks. Subsequently, forty patients scheduled for CABG were randomized into two groups, receiving, respectively, a) standard medical and surgical treatment (20 patients) and b) the same treatment plus 40,000 IU of EPOalpha in a single bolus injection in the immediate pre-surgical period (20 patients). In our population, we did not find any correlation between pre-surgical EPO and post-surgical troponin I or CK-MB peaks (p Pearson > 0.05). Furthermore, patients treated with EPOalpha did not show differences compared to the control group in either troponin I (1.7+/-1.8 vs 2.6+/-3.4, p>0.05) or CK-MB (19.6 +/-13.2 vs 17.1+/-12.6, p>0.05) peaks measured in the post-surgical period.

Topics: Aged; Cardiopulmonary Bypass; Coronary Artery Bypass, Off-Pump; Creatine Kinase, MB Form; Epoetin Alfa; Erythropoietin; Female; Humans; Male; Middle Aged; Recombinant Proteins; Reperfusion Injury; Troponin I

Ovarian torsion is one of the most dangerous gynecological emergencies requiring surgery. A total of 50%-90% ovarian torsion cases are caused by physiological cysts, endometriosis, and other benign or malignant ovarian neoplasms. The aim of the study was to investigate the effects of erythropoietin (EPO) treatment on ischemia/reperfusion (IR) injury caused by ovarian torsion/detorsion (T/D) injury. Thirty female Wistar albino rats were divided into five groups as follows: Group I: Control; Group II: Torsion (T); Group III: Torsion/Detorsion(T/D); Group IV: Torsion/Detorsion (T/D) + EPO; Group V: EPO. Sections of the ovaries were evaluated for histopathological changes with hematoxylin and eosin stain, a immunohistochemical assay for caspase 3 expression, and the TUNEL assay for apoptosis. Ovarian sections from torsion/detorsion and torsion groups showed more hemorrhage, vascular congestion, edema, degenerative granulosa, and stromal cells. Fewer histopathological changes were found in EPO and T/D + EPO groups. Caspase 3 and TUNEL positive cells were significantly increased in the torsion/detorsion group as compared with the other groups (

Topics: Animals; Antioxidants; Caspase 3; Epoetin Alfa; Erythropoietin; Female; Humans; Ischemia; Ovarian Diseases; Ovarian Torsion; Rats; Rats, Wistar; Reperfusion Injury; Torsion Abnormality

Skeletal muscle ischemia and reperfusion (S-I/R) injury is relieved by interventions like remote ischemic preconditioning (RIPC). Here, we tested the hypothesis that simultaneous exposure to a minimal dose of erythropoietin (EPO) boosts the protection conferred by RIPC against S-I/R injury and concomitant mitochondrial oxidative and apoptotic defects.. S-I/R injury was induced in rats by 3-h right hindlimb ischemia followed by 3-h of reperfusion, whereas RIPC involved 3 brief consecutive I/R cycles of the contralateral hindlimb.. S-I/R injury caused (i) rises in serum lactate dehydrogenase and creatine kinase and falls in serum pyruvate, (ii) structural deformities like sarcoplasm vacuolations, segmental necrosis, and inflammatory cells infiltration, and (iii) decreased amplitude and increased duration of electromyography action potentials. These defects were partially ameliorated by RIPC and dose-dependently by EPO (500 or 5000 IU/kg). Further, greater repairs of S-I/R-evoked damages were seen after prior exposure to the combined RIPC/EPO-500 intervention. The latter also caused more effective (i) preservation of mitochondrial number (confocal microscopy assessed Mitotracker red staining) and function (citrate synthase activity), (ii) suppression of mitochondrial DNA damage and indices of oxidative stress and apoptosis (succinate dehydrogenase, myeloperoxidase, cardiolipin, and cytochrome c), (iii) preventing calcium and nitric oxide metabolites (NOx) accumulation and glycogen consumption, and (iv) upregulating EPO receptors (EPO-R) gene expression.. dual RIPC/EPO conditioning exceptionally mends structural, functional, and neuronal deficits caused by I/R injury and interrelated mitochondrial oxidative and apoptotic damage. Clinically, the utilization of relatively low EPO doses could minimize the hormone-related adverse effects.

Topics: Animals; Erythropoietin; Ischemia; Ischemic Preconditioning; Muscle, Skeletal; Rats; Reperfusion Injury

Several studies provide evidence that erythropoietin (EPO) could play an important role in the recovery of the heart subjected to ischemia-reperfusion. In this regard, it has been suggested that EPO could be involved in protein kinase B (Akt) activation as a cell survival protein. The aim of the present study was to investigate the effects of EPO on the Akt/glycogen synthase kinase 3 beta (GSK-3β) pathway in the presence or absence of wortmannin (W, Akt inhibitor) and its relationship with mitochondrial morphology and function preservation in ischemic-reperfused rat hearts. EPO improved the functional recovery of the heart subjected to ischemia-reperfusion, reduced the release of CK and the infarct size, and promoted preservation of the mitochondrial structure. Moreover, it reduced tissue lactate content and preserved glycogen in order to prevent ischemia. The results showed greater Akt activation, accompanied by preservation of swelling and mitochondrial calcium retention capacity, as well as an increase in ATP synthesis capacity. These results were accompanied by an inhibition of GSK-3β, suggesting regulation of Akt on the opening of the mitochondrial permeability transition pore. All these beneficial effects exerted by acute treatment with EPO were prevented by W. The present study provided novel evidence that EPO not only enhances intrinsic activation of Akt during myocardial ischemia-reperfusion but also promotes GSK-3β inhibition, contributing to mitochondrial structure and function preservation.

Topics: Animals; Cardiotonic Agents; Erythropoietin; Glycogen Synthase Kinase 3 beta; Heart; Ischemia; Phosphorylation; Proto-Oncogene Proteins c-akt; Rats; Reperfusion Injury

To explore the effect of erythropoietin (EPO) on the AMP-activated protein kinase (AMPK)/nicotinamide adenine dinucleotide phosphatase oxidase 4 (NOX4) signaling pathway during renal ischemia reperfusion injury (RIRI) in rats.. A rat model of RIRI was established by clamping the left renal pedicle and removing the right kidney. The rats in the sham group did not have their left renal pedicle clamped. Rats with a model of RIRI were randomly divided into RIRI alone (control), erythropoietin treatment (EPO/RIRI), and Compound C treatment (CPC/RIRI) groups. Hematoxylin-eosin (H&E) staining was used to examine pathological kidney damage. Serum creatinine and urea nitrogen levels were measured to evaluate renal function. Western blotting was performed to detect the expression levels of phosphorylated p-AMPK and total AMPK protein in the kidneys. RT-PCR was used to evaluate the mRNA levels of Nox4 and p22 in the kidneys. Oxidative stress-related indices (ROS, CAT, GSH, SOD, and MDA) were also measured.. EPO treatment improved kidney function by preventing kidney damage induced by the RIRI model. Preventing ischemia/reperfusion injury in the RIRI model was correlated with an increased p-AMPK/AMPK ratio and elevated activity of CAT, GSH, and SOD, which ameliorated the expression of NOX4, p22, ROS, and MDA. Moreover, treatment with CPC (an AMPK inhibitor) reduced the effects of EPO in the RIRI model.. EPO treatment protected rats against RIRI in the RIRI model by alleviating oxidative stress by triggering the AMPK/NOX4/ROS pathway.

Topics: AMP-Activated Protein Kinases; Animals; Erythropoietin; Kidney; NADPH Oxidase 4; Oxidative Stress; Rats; Reactive Oxygen Species; Reperfusion Injury; Superoxide Dismutase

Ischemia reperfusion (IR) injury of skeletal muscles is a serious problem because of its local and systemic complications. Previous studies reported that ozone and erythropoietin could alleviate IR effect on several organs. The current research is established to evaluate the possible protective role of ozone versus erythropoietin following IR injury of the gastrocnemius muscle. Fifty rats were equally divided into five groups: I control, II ischemia reperfusion (IR), III post-reperfusion ozone treated, IV post-reperfusion erythropoietin-treated, and V recovering post-reperfusion without treatment groups. The right femoral arteries of all rats were clamped for three hours to induce ischemia then clamps were released to allow reperfusion for two hours. Rats of group II were scarified immediately after reperfusion period. Rats of group III were injected with ozone just after reperfusion for 14 days. Animals of group IV were injected with erythropoietin just after reperfusion for 14 days. Rats of group V rats were kept for 2 weeks following reperfusion without treatment. Blood samples were obtained to estimate lactate dehydrogenase (LDH) and creatine kinase (CK) enzymes. Gastrocnemius muscle was processed for measurement of tissue malondialdehyde (MDA), as well as examination by light and electron microscopes. iNOS and PCNA immunohistochemistry and statistical analysis were applied. The current results indicated that both ozone and erythropoietin could be used as protective agents reducing the muscular damage induced by IR injury.

Topics: Animals; Erythropoietin; Male; Muscle, Skeletal; Ozone; Rats; Reperfusion Injury

Renal ischemia/reperfusion (RI/R) is a critical clinical outcome with slightly reported improvement in mortality and morbidity. Effective therapies are still crucially required. Accordingly, the therapeutic effects of esculin (ESC, LCESI-MS/MS-isolated compound from Vachellia farnesiana flowers extract, with reported P2X7 receptor inhibitor activity) alone and in combination with erythropoietin (EPO) were investigated against RI/R injury and the possible underlying mechanisms were delineated. ESC and EPO were administered for 7 days and 30 min prior to RI, respectively. Twenty-four hour following reperfusion, blood and kidney samples were collected. Results revealed that pretreatment with either ESC or EPO reduced serum nephrotoxicity indices, renal oxidative stress, inflammatory, and apoptosis markers. They also ameliorated the renal histopathological injury on both endothelial and tubular epithelial levels. Notably, ESC markedly inhibited P2X7 receptors and NLRP3 inflammasome signaling (downregulated NLRP3 and Caspase-1 gene expressions), whereas EPO significantly upregulated PI3K and Akt gene expressions, also p-PI3K and p-Akt levels in renal tissues. ESC, for the first time, demonstrated effective protection against RI/R-injury and its combination with EPO exerted maximal renoprotection when compared to each monotherapy, thereby representing an effective therapeutic approach via inhibiting oxidative stress, inflammation, renal tubular and endothelial injury, apoptosis, and P2X7 receptors expression, and activating PI3K/Akt pathway.

Topics: Apoptosis; Erythropoietin; Esculin; Humans; Kidney; NLR Family, Pyrin Domain-Containing 3 Protein; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Receptors, Purinergic P2X7; Reperfusion Injury; Tandem Mass Spectrometry

Topics: Animals; Epoetin Alfa; Erythropoietin; Female; Humans; Rats; Rats, Wistar; Recombinant Proteins; Relaxin; Reperfusion Injury; Superoxide Dismutase; Uterus

Although electroacupuncture (EA) has been shown to be effective in the treatment of stroke, its mechanisms of action remain undefined. This study explored the therapeutic effects of EA in rats with cerebral ischemia-reperfusion injury (CIRI) and evaluated its possible mechanisms in promoting angiogenesis. To evaluate the effect of EA, we used 2, 3, 5-Triphenyl-2H-Tetrazolium Chloride (TTC) staining and behavior score to calculate the cerebral infarct volume and neurological deficit score after CIRI. Western blot (WB) analysis was employed to evaluate the expression of cluster of differentiation 34 (CD34), erythropoietin (EPO), vascular endothelial growth factor (VEGF) and phospho-Src (p-Src) in the brain of the rats with CIRI. On the other hand, we established an oxygen-glucose deprivation/reoxygenation (OGD/R) injury model using brain microvascular endothelial cells (BMECs), and analyzed cell viability and expression of VEGF or p-Src using cell counting kit-8 (CCK-8) and WB, respectively. Our data showed that EA at the GV26 acupoint could significantly promote the expression of CD34, EPO, VEGF and p-Src in CIRI rats. Our CCK-8 results demonstrated that intervention with recombinant EPO and VEGF proteins remarkably improved the viability of BMECs after OGD/R, while a Src inhibitor, PP1, reversed this phenotype. The WB results showed that the recombinant EPO protein increased the expression of VEGF and p-Src, which was significantly inhibited by PP1. Taken together, our findings showed that EA at the GV26 acupoint can significantly attenuate ischemic injury after stroke and promote angiogenesis via activation of EPO-mediated Src and VEGF signaling pathways. Besides, the upregulation of VEGF may also be associated with the activation of Src by EPO.

Topics: Animals; Chlorides; Electroacupuncture; Endothelial Cells; Erythropoietin; Glucose; Ischemia; Oxygen; Rats; Reperfusion Injury; Signal Transduction; Stroke; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

The aim: This study aimed to examine the anti-inflammatory, and antiapoptotic effects of erythropoietin against kidney injury inducted by ischemia reperfusion in experimental model.. Materials and methods: 20 male Sprague Dawley rats were randomly divided into 4 equal groups: sham (subject to median laparotomy only), control (subject to 30 minutes ischemia and 2hours reperfusion), vehicle (injected by distilled water and subjected to the same procedure of ischemia reperfusion), erythropoietin group (as in vehicle group but the rats pretreated with 1000 U/kg of erythropoietin). The left kidney and blood specimen were collected. The blood utilized to assess serum creatinine. While kidneys utilized to assessed MCP-1, TLR2, and caspase-3 in addition to histopathological evaluation.. Results: Control and vehicle samples showed that a significant elevation in serum creatinine, TLR2, caspase-3, and MCP-1 as compared with sham group. The histological eval¬uation showed a significant rise in kidney injury scores. Kidneys and blood samples of erythropoietin pretreated rats established histopathological and functional improvement as evidenced via reduced kidney injury scores in addition to the reduction in serum creatinine, as well as there were a significant diminished in caspase-3, MCP-1, and TLR2 levels when compared with control and vehicle groups.. Conclusions: Erythropoietin has renoprotective effect against ischemia and reperfusion, which achieved by decrease the inflammatory response as well as antiapoptotic effect.

Topics: Animals; Apoptosis; Caspase 3; Creatinine; Erythropoietin; Ischemia; Kidney; Male; Rats; Rats, Sprague-Dawley; Reperfusion; Reperfusion Injury; Toll-Like Receptor 2

Undescended testis is an absence of testis in the scrotum, the incidence was 15 cases per 1000 from 1974 to 1996 in Europe. At Saiful Anwar Regional Hospital East Java, from January 2015 to July 2019 there were 60 boys diagnosed with undescended testis. A temperature rise of testis located in the abdominal triggers production of reactive oxygen species, causing impairment of the testicular epithelial germ cells and spermatogenesis, leading to many complications. Erythropoietin is a glycoprotein hormone that circulates in the body and has a positive effect on ischemic injury/gonadal reperfusion.. To find out ROS involvement in undescended testis and efficacy of EPO as an additional therapy for undescended testis.. This study is an experimental study with a post-test only control group design, using 18 male Wistar mice conditioned to be undescended testis for 7 days and underwent orchidopexy and some are given additional erythropoietin 1000iu/Kg 3 times a week.. Before and after the intervention, the mean body weight of mice did not experience a significant difference, meanwhile testicular volume showed a significant difference between the orchidopexy and EPO groups (p = 0.005 and 0.001). Johnsen's score were found significant in the EPO group. Malone dialdehyde level in EPO and orchidopexy group showed significant difference p = 0.01 and 0.009 when compared to undescended testis group.. There was the involvement of ROS in undescended testis and additional EPO improve impairment of germinal epithelial cells and spermatogenesis process due to undescended testis.

Topics: Animals; Cryptorchidism; Epithelium; Erythropoietin; Humans; Male; Mice; Reperfusion Injury; Testis

Topics: Acute Lung Injury; Animals; Cell Line; Erythropoietin; Humans; p38 Mitogen-Activated Protein Kinases; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction

Chronic kidney disease (CKD) or acute kidney injury (AKI) causes impaired kidney function, leading to cognitive impairment, neuropathy, and cerebrovascular disease. Due to kidney damage, toxins stay in the blood rather than leaving the body through the urine, and brain function is affected by kidney-brain interaction. The present study aimed to investigate the protective effects of erythropoietin mimetic peptide (pHBSP) and infliximab on ischemic renal reperfusion injury. The experiment was performed on 70 white male Wistar laboratory rats which received recombinant erythropoietin, pHBSP, and infliximab. Under anesthesia, traumatic vascular clamps were applied to the left renal pedicle for 40 min, and nephrectomy was performed on the right. Functional tests and laboratory tests were performed 5 min and 24 h after the reperfusion.  Thereafter, 24 h after the surgery, the plasma creatinine and urea levels in the sham-operated animals were obtained at 45.9±0.8 mmol/L and 6.7±0.2 mmol/L, respectively. Plasma creatinine and urea levels in the control group animals were 102.63±3.6 mmol/L and 21.80±1.29 mmol/L, respectively. The administration of pHBSP and infliximab to the animals with ischemia-reperfusion kidney injury has a pronounced nephroprotective effect, as compared to erythropoietin. There was a significant decrease in blood levels of creatinine and urea, improvement of microcirculation in the kidney, normalization of glomerular filtration rate, and fractional sodium excretion. The results of the study demonstrated pointed to the prospects of pHBSP and infliximab administration in ischemia-reperfusion kidney injury and justified the feasibility of further research in this field.

Topics: Animals; Erythropoietin; Infliximab; Kidney; Male; Rats; Rats, Wistar; Reperfusion Injury

Previous studies have indicated that EPO maintains the M2 microglia phenotype that contributes to white matter repair after ischemic stroke in young mice (2 months old). However, the underlying mechanisms that regulate microglial polarization are poorly defined. This study investigated the neuroprotective effects of nonerythropoietic mutant EPO (MEPO) on white matter and the underlying mechanism in middle-aged (9-month-old) male mice following cerebral ischemia. Middle-aged male C57 BL/6 mice were treated with MEPO (5000 IU/kg) or vehicle after middle cerebral artery occlusion (MCAO) and reperfusion. The specific inhibitor AG490 was used to block the JAK2/STAT3 pathway. Neurological function was assessed by beam walking and adhesive removal tests. Immunofluorescence staining and western blotting were used to assess the severity of white matter injury, phenotypic changes in the microglia and the expression of the signaling molecules. MEPO significantly improved neurobehavioral outcomes, alleviated brain tissue loss, and ameliorated white matter injury after MCAO compared with the vehicle group. Moreover, MEPO promoted oligodendrogenesis by shifting microglia toward M2 polarization by promoting JAK2/STAT3 activation and inhibiting the expression of C/EBPβ at 14 days after cerebral ischemia-reperfusion. However, the MEPO's effect on microglial M2 polarization and oligodendrogenesis was largely suppressed by AG490 treatment. Collectively, these data indicate that MEPO treatment improves white matter integrity after cerebral ischemia, which may be partly explained by MEPO facilitating microglia toward the beneficial M2 phenotype to promote oligodendrogenesis via JAK2/STAT3 and the C/EBPβ signaling pathway. This study provides novel insight into MEPO treatment for ischemic stroke.

Topics: Animals; Behavior, Animal; Brain Ischemia; CCAAT-Enhancer-Binding Protein-beta; Cell Polarity; Erythropoietin; Janus Kinase 2; Male; Mice; Mice, Inbred C57BL; Microglia; Mutation; Oligodendroglia; Recovery of Function; Reperfusion Injury; Signal Transduction; STAT3 Transcription Factor; Steroid Isomerases; Tyrphostins; White Matter

Ischemic preconditioned (IP) neurons protect astrocytes against ischemia/reperfusion (I/R)-induced injury by inhibiting oxidative stress. However, the relevant mechanisms are unknown. Based on the role of nuclear factor-κB (NF-κB) in cell survival and adaption to oxidative stress, we hypothesized that NF-κB might be associated with astroprotection induced by IP neurons via upregulation of antioxidant enzymes. Here, we investigated the effects of IP neurons on NF-κB activation, cell viability, reactive oxygen species (ROS), expression of antioxidant enzymes, erythropoietin (EPO), and tumor necrosis factor α (TNF-α), in the presence or absence of BAY11-7082 (an NF-κB inhibitor), anti-EPO, and anti-TNF-α antibodies, in astrocytes treated with or without I/R. We found that IP neurons could keep NF-κB activation at a relatively higher but beneficial level, and in turn, upregulated the activity of antioxidant enzymes and hence enhanced cell viability and reduced ROS in I/R treated astrocytes. The results collectively indicated that IP neurons are able to significantly inhibit the I/R-induced NF-κB overactivation, probably via EPO and TNF-α, being essential for IP neuron-induced astroprotection under the conditions of I/R. We concluded that NF-κB-mediated antioxidative stress is one of the mechanisms by which IP neurons protect astrocytes against I/R injury.

Topics: Animals; Antioxidants; Astrocytes; Cell Hypoxia; Cells, Cultured; Cerebral Cortex; Culture Media, Conditioned; Erythropoietin; Glucose; Neurons; NF-kappa B; Oxidative Stress; Paracrine Communication; Rats, Sprague-Dawley; Reactive Oxygen Species; Reperfusion Injury; Signal Transduction; Tumor Necrosis Factor-alpha

Renal ischemia-reperfusion (IR) injury and cyclosporine A (CsA) nephrotoxicity affect allograft function and survival. The prolonged effects and underlying mechanisms of erythropoietin derived cyclic helix B peptide (CHBP) and/or caspase-3 small interfering RNA (CASP-3siRNA) were investigated in mouse kidneys, as well as kidney epithelial cells (TCMK-1), subjected to transplant-related injuries. Bilateral renal pedicles were clamped for 30 min followed by reperfusion for 2 and 8 weeks, with/without 35 mg/kg CsA gavage daily and/or 24 nmol/kg CHBP intraperitoneal injection every 3 days. The ratio of urinary albumin to creatinine was raised by IR injury, further increased by CsA and lowered by CHBP at 2, 4, 6 and 8 weeks, whereas the level of SCr was not significantly affected. Similar change trends were revealed in tubulointerstitial damage and fibrosis, HMGB1 and active CASP-3 protein. Increased apoptotic cells in IR kidneys were decreased by CsA and CHBP at 2 and/or 8 weeks. p70 S6 kinase and mTOR were reduced by CsA with/without CHBP at 2 weeks, so were S6 ribosomal protein and GSK-3β at 8 weeks, with reduced CASP-3 at both time points. CASP-3 was further decreased by CHBP in IR or IR + CsA kidneys at 2 or 8 weeks. Furthermore, in TCMK-1 cells CsA induced apoptosis was decreased by CHBP and/or CASP-3siRNA treatment. Taken together, CHBP predominantly protects kidneys against IR injury at 2 weeks and/or CsA nephrotoxicity at 8 weeks, with different underlying mechanisms. Urinary albumin/creatinine is a good biomarker in monitoring the progression of transplant-related injuries. CsA divergently affects apoptosis in kidneys and cultured kidney epithelial cells, in which CHBP and/or CASP-3siRNA reduces inflammation and apoptosis.

Topics: Animals; Apoptosis; Caspase 3; Caspase Inhibitors; Cell Line; Cyclosporine; Disease Models, Animal; Erythropoietin; Kidney; Macrophages; Male; Mice; Mice, Inbred BALB C; Peptide Fragments; Peptides, Cyclic; Reperfusion Injury; RNA, Messenger; RNA, Small Interfering

Hypoxia‑inducible factor‑1α (HIF‑1α) is a key transcriptional factor in response to hypoxia and is involved in ischemic stroke. In the present study, the potential for HIF‑1α to inhibit neuronal apoptosis through upregulating erythropoietin (EPO) was investigated in a transient middle cerebral artery occlusion (tMCAO) rat stroke model. For this purpose, a recombinant adenovirus expressing HIF‑1α was engineered (Ad‑HIF‑1α). Control adenovirus (Ad group), Ad‑HIF‑1α (Ad‑HIF‑1α group) or Ad‑HIF‑1α in addition to erythropoietin mimetic peptide‑9 (EMP9), an EPO‑receptor (‑R) antagonist (Ad‑HIF‑1α+EMP9 group), were used for an intracranial injection into rat ischemic penumbra 1 h following MCAO. All rats demonstrated functional improvement following tMCAO, while the improvement rate was faster in rats treated by Ad‑HIF‑1α compared with all other groups. The EPO‑R inhibitor partially reversed the benefits of Ad‑HIF‑1α. Apoptosis induced by tMCAO was significantly inhibited by Ad‑HIF‑1α (P<0.05). The expression of HIF‑1α, evaluated by immunohistochemistry either in neurons or astrocytes, was upregulated by Ad‑HIF‑1α. Both EPO mRNA and protein expression were increased by Ad‑HIF‑1α, however, there was no significant change of EPO‑R either on an mRNA level or protein level. Furthermore, EMP9 did not change the EPO expression which was upregulated by Ad‑HIF‑1α. Activated caspase 3 in neurons was suppressed by Ad‑HIF‑1α. Activated caspase 3 downregulated by HIF‑1α was partially blocked by EMP9. Altogether, the present data demonstrated that HIF‑1α attenuates neuronal apoptosis partially through upregulating EPO following cerebral ischemia in rat. Thus, upregulating HIF‑1α subsequent to a stroke may be a potential treatment for ischemic stroke.

Topics: Animals; Apoptosis; Brain Diseases; Disease Models, Animal; Erythropoietin; Gene Expression Regulation; Hypoxia-Inducible Factor 1, alpha Subunit; Male; Neurons; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Acute kidney injury (AKI) is the most common condition in hospitalized patients. As ischemia/reperfusion-induced AKI (IR-AKI) is as a major contributor to end-stage disease, an effective therapeutic intervention for IR-AKI is imperative. Erythropoietin (EPO) is a potent stimulator of erythroid progenitor cells and is significantly upregulated during hypoxia. Here, we investigated the renoprotective effects of EPO in an IR-AKI mouse model. Mice were assigned to sham, EPO only, and IR only groups, and the IR group was treated with EPO prior to injury. EPO was administered twice at 30 min prior to bilateral renal artery occlusion, and 5 min before reperfusion, with all mice sacrificed 24 h after IR-AKI. The serum was harvested for renal functional measurements. The kidneys were subjected to histological evaluation, and the biochemical changes associated with renal injury were assessed. EPO significantly attenuated the renal dysfunction associated with IR-AKI, as well as tissue injury. Apoptotic cell death and oxidative stress were significantly reduced in EPO-treated mice. Macrophage infiltration and expression of ICAM-1 and MCP-1 were also significantly reduced in EPO-treated mice. Furthermore, the expression of inflammasome-related factors (NLRP1, NLRP3, and caspase-1 cleavage), via the activation of the COX-2 and NF-B signaling pathways were significantly reduced following EPO treatment. To our knowledge, this is the first study to demonstrate that inflammasome-mediated inflammation might be a potential target of EPO as a treatment for ischemic AKI.

Topics: Acute Kidney Injury; Adaptor Proteins, Signal Transducing; Animals; Apoptosis; Apoptosis Regulatory Proteins; Caspase 1; Cell Hypoxia; Cyclooxygenase 2; Disease Models, Animal; Erythroid Precursor Cells; Erythropoietin; Gene Expression Regulation; Humans; Inflammasomes; Kidney; Mice; NLR Family, Pyrin Domain-Containing 3 Protein; Reperfusion Injury

The purpose of this study was to examine the role of endothelial progenitor cells (EPCs) in protection against ischemic-reperfusion injury (IRI) in a nephron-sparing surgery (NSS) rat model using erythropoietin (EPO) preconditioning. Fifty-four male Sprague-Dawley rats were randomly divided into 3 groups for right kidney nephrectomy treatment: sham group (exposure without clamp treatment), NSS group (3 days of peritoneal phosphate buffered saline [PBS] injection before renal blood vessels were clamped for 40 mins and NSS was performed), and EPO group (3 days of EPO abdomen injections prior to renal blood vessel clamping for 40 min before NSS was performed). After 12, 24, and 72 hours, inferior vena cava blood and renal tissues were harvested. The extent of renal injury was assessed, along with EPC number, cell proliferation, angiogenesis, and vascular growth factor expression. EPO preconditioning significantly improved renal function and histologic morphology, indicated by reduced blood urea nitrogen (BUN) ([33.12 ± 1.88] vs [16.03 ± 0.91], P < .05) and serum creatinine (Scr) ([190.2 ± 20.23] vs [77.23 ± 5.82], P < .05) levels and histologic injury scores ([3.20 ± 0.78] vs [1.70 ± 0.67], P < .05). Angiogenesis in peritubular capillaries markedly increased in the EPO group. EPC numbers increased in the kidneys at 24 hours following reperfusion in the EPO group, compared to the NSS group. Furthermore, EPO preconditioning also increased SDF-1α and CXCR7 expression at 24 hours following reperfusion relative to the NSS group. These findings suggest that EPO pretreatment can reduce renal injury in rats caused by IRI. Mechanistically, this may be related to EPC mobilization and recruitment to injured renal tissues by SDF-1α and CXCR7.

Topics: Animals; Endothelial Progenitor Cells; Erythropoietin; Kidney; Male; Nephrons; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Urologic Surgical Procedures

Topics: Animals; Apoptosis; Caspase 3; Creatinine; Disease Models, Animal; Drug Therapy, Combination; Erythropoietin; HMGB1 Protein; Injections, Intraperitoneal; Injections, Intravenous; Kidney; Male; Mice; Mice, Inbred C57BL; Peptide Fragments; Reperfusion Injury; RNA, Small Interfering; Transcriptome

Kidney ischemia reperfusion (IR) contributes to the development of acute kidney injury. The hypoxic conditions in ischemic damage lead to oxidative stress and apoptotic cell death. We investigated the effects of vitamin D3 (Vit D) and erythropoietin (EPO) on microRNA-21(miR-21) expression in renal IR. Wistar rats were divided into five groups including the control, vehicle + IR, Vit D + IR, EPO + IR, and Vit D + EPO + IR groups. The animals were unilaterally nephrectomized and subjected to 45 min of renal pedicle occlusion followed by 24 h reperfusion. Vitamin D3 and EPO were administered prior to ischemia. After 24 h reperfusion, the kidney samples were collected for the detection of miR-21, heat shock protein 70 (hsp70) and caspase-3 expression levels. Kidney IR significantly increased the expression of miR-21, hsp70 and capase-3 and blood urea nitrogen (BUN)-Cr levels. Treatment with vitamin D3 and EPO significantly decreased the BUN-Cr levels and hsp70 and caspase-3 expression. Also, the co-administration of two drugs significantly increased miR-21 expression. It seems that vitamin D3 or EPO administration could protect the kidney against IR injury. However, vitamin D3 and EPO co-treatment was the most effective compared with the other treatment groups.

Topics: Animals; Blood Urea Nitrogen; Cholecalciferol; Erythropoietin; HSP70 Heat-Shock Proteins; Kidney; Kidney Glomerulus; Kidney Tubules; MicroRNAs; Rats; Rats, Wistar; Reperfusion Injury

Substantial progress was made in acute kidney injury (AKI) over the past 10 years, but no therapeutic interventions have been shown to prevent AKI or accelerate functional recovery after injury. A large number of preclinical studies supports the use of recombinant human erythropoietin (rHuEPO) to prevent AKI, but the clinical trial data are inconclusive. To address concerns about preclinical study design and reporting in AKI, we here presented our rigorous experiments on the use of rHuEPO in a mouse model simulating the most common post-ischemic AKI in patients.. Use of saline vehicle or rHuEPO (100 or 1000 U/KgBW) in mice subjected to AKI induced by ischemia-reperfusion injury of left kidney 2 weeks after right nephrectomy (NX + IRI).. NX + IRI resulted in a reproducible AKI model. Use of rHuEPO as a pretreatment or posttreatment did not affect AKI severity, functional recovery, and mouse survival regardless of gender, injury severity, or doses of rHuEPO. Administering rHuEPO with 1000 U/KgBW did increase hematocrit and modulate AKI kidney macrophages by Nos2 downregulation and Ccl17 upregulation. Active expression of erythropoietin receptor (EPOR) was not identified in renal cells by lineage tracing study, whereas expression of colony-stimulating factor 2 receptor β (CSF2Rβ) was identified in kidney macrophages and upregulated after AKI. Both EPOR and CSF2Rβ were identified in cultured bone marrow derived macrophages, possibly mediated the robust inhibition of cytokine-induced phenotype switching by rHuEPO.. Use of rHuEPO can modulate macrophage function but not the post-ischemic AKI severity, functional recovery and survival in mice.

Topics: Acute Kidney Injury; Animals; Apoptosis; Disease Models, Animal; Erythropoietin; Female; Kidney; Macrophages; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Nephrectomy; Recombinant Proteins; Reperfusion Injury; Signal Transduction

Kidney ischemia and reperfusion injury could cause microvascular barrier dysfunction, lung inflammatory cascades activation, and programmed cell death of pulmonary endothelium, leading to acute lung injury. Our study aimed at determining whether erythropoietin (EPO) can ameliorate lung dysfunction following renal ischemia and reperfusion (IR) injury and explored the underlying mechanisms.. In vivo, C57BL/6 mice received EPO (6000 U/kg) before right renal vascular pedicles clamping for 30 minutes, followed by 24 hours of reperfusion. The lung histopathologic changes and inflammatory cytokines expression were assessed. In vitro, cultured human umbilical vein endothelial cells were treated with EPO, and apoptosis rate, proliferation capacity, and phosphorylation status of the Janus kinase-signal transducer and activator of transcription 3 (Jak-STAT3) pathway were measured respectively in the presence or absence of lipopolysaccharide stimulation.. In vivo, EPO remarkably attenuated pulmonary interstitial and alveolar epithelial edema caused by renal IR injury. In vitro, the proliferation capacity of human umbilical vein endothelial cells was significantly increased under EPO stimulation, which correlated with changes in Jak-STAT3 signaling.. Our data indicated that EPO is able to ameliorate acute lung tissue damage induced by renal IR, and at least in part, via the Jak-STAT3 pathway.

Topics: Acute Lung Injury; Animals; Cell Line; Cell Proliferation; Endothelium, Vascular; Erythropoietin; Humans; Janus Kinases; Kidney; Kidney Diseases; Male; Mice; Mice, Inbred C57BL; Reperfusion Injury; Signal Transduction; STAT3 Transcription Factor

Since the nature of ischemia/reperfusion (IR)-induced tissue damage is multifactorial and complex, in the current study, the effects of multiple treatment strategies via concomitant administration of erythropoietin (EPO) and N-acetylcysteine (NAC) with an ischemic preconditioning (IPC) regimen on renal IR injury were examined.. Thirty male Wistar rats were subjected to bilateral occlusion of the renal pedicles for 50 min followed by reperfusion. EPO (1000 IU/kg) was administered for 3 days, as well as IPC before the IR and NAC (150 mg/kg) administration for 4 days after IR. The animals were randomly allocated into 6 groups (n = 5): sham, IR, EPO+IR, IPC+IR, NAC+IR, and EPO+IPC+NAC+IR. Kidney tissues and blood samples were obtained for oxidative stress, proinflammatory cytokines, and renal functional evaluations.. IR caused significant inflammatory response, oxidative stress, and reduced renal function. Treatment with EPO, IPC, and NAC or a combination of two of them attenuated renal dysfunction and reduced the oxidative stress and inflammatory markers. Rats treated with the combination of EPO, IPC, and NAC showed a higher degree of protection compared to the other groups.. These results showed that concomitant administration of EPO and IPC along with posttreatment NAC may have additive beneficial effects on kidney IR injury during IR-induced acute renal failure.

Topics: Acetylcysteine; Animals; Blood Urea Nitrogen; Creatinine; Cytokines; Disease Models, Animal; Erythropoietin; Ischemic Preconditioning; Kidney; Kidney Diseases; Male; Oxidative Stress; Rats; Rats, Wistar; Reperfusion Injury

Condensation Erythropoietin improved the survival of follicles in ovarian grafts most likely by reducing ischemic injury, by improving neoangiogenesis, and by its antioxidant effects.. Ovarian tissue cryopreservation and transplantation are the only options accepted for prepubertal girls and women requiring immediate chemotherapy. Ischemia-reperfusion injury is the main obstacle for ovarian tissue transplantation. In the present study, we aimed to evaluate the effects of recombinant human erythropoietin (EPO) on tissue viability in autotransplanted rat ovaries.. Seventeen female rats were randomized into 3 groups as sham control group (n = 5), EPO-treated group (n = 6), and EPO-untreated group (n = 6). Both ovaries were excised and transplanted into a subcutaneous pouch formed at the anterior abdominal wall in the EPO-treated and untreated groups. In the EPO group, 5000 U/kg EPO was applied as local injection to the site that ovarian tissue was placed and the dose was repeated with the same route at the end of the fourth week. After 2 months, ovaries were removed and blood samples were obtained. Levels of estradiol (E. Considering serum levels, mean CAT was significantly higher ( P = .003) and mean SOD ( P = .033), LPO ( P = .050), VEGF ( P = .001), and VEGF-C ( P = .024) were significantly lower in the EPO-treated group than in the untreated group. Mean serum GPX levels were similar. Significantly higher levels of E. Erythropoietin improved the survival of follicles in ovarian grafts most likely by reducing ischemic injury, by improving neoangiogenesis, and by its antioxidant effects.

Topics: Animals; Antioxidants; Catalase; Erythropoietin; Estradiol; Female; Neovascularization, Physiologic; Ovary; Oxidative Stress; Rats; Rats, Wistar; Reperfusion Injury; Superoxide Dismutase; Vascular Endothelial Growth Factor A

Ischemia/reperfusion injury (IRI) commonly occurs in renal transplantation. Erythropoietin (EPO) exerts a protective effect in IRI. To investigate the underlying molecular mechanism, rat models of renal IRI were established and treated with EPO and/or lentivirus‑mediated EPO-siRNA, the signal transducer and activator of transcription 6 (STAT6) inhibitor AS1517499, the JNK inhibitor SP600125, the p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580, and the nuclear factor (NF)-κB inhibitor lactacystin. Histological examination revealed that EPO protected the kidney from IRI, through decreasing the extent of tissue congestion and inflammatory cell infiltration; however, EPO siRNA did not exert the same protective effect. In addition, the EPO level was inversely associated with renal IRI. EPO downregulated the expression of interferon-γ, interleukin (IL)-4, creatinine and caspase-3, and upregulated the expression of IL-10, thymic stromal lymphopoietin, STAT6, p-JNK and p-p38, while the opposite effects were observed with the administration of EPO-siRNA and the specific respective inhibitors. Further results revealed that MAPK (p-JNK and p-p38) acted upstream of NF-κB, and that NF-κB signaling regulated the expression of caspase-1 and -3, which may be responsible for the cytotoxicity associated with IRI. Taken together, the results of the present study demonstrated that EPO exerted a protective effect in renal IRI via the STAT6/MAPK/NF-κB pathway. This protective effect of EPO may improve reperfusion tolerance in ischemic kidneys and benefit transplant recipients.

Topics: Acetylcysteine; Animals; Anthracenes; Apoptosis; Erythropoietin; Gene Expression Regulation; Humans; Imidazoles; Interferon-gamma; Kidney; Kidney Transplantation; Lentivirus; MAP Kinase Kinase 4; NF-kappa B; p38 Mitogen-Activated Protein Kinases; Pyridines; Pyrimidines; Rats; Reperfusion Injury; RNA, Small Interfering; Signal Transduction; STAT6 Transcription Factor

BACKGROUND Lung ischemia/reperfusion injury (LIRI) is a medical problem featuring pulmonary dysfunction and damage. The present study aimed to investigate the protective effects of erythropoietin (EPO), which has been reported to be an anti-inflammatory agent, on LIRI through inhibiting the TLR-4/NF-κB signaling pathway. MATERIAL AND METHODS All rats were randomly divided into 3 groups (n=8): a control group, a vehicle+LIRI group, and an EPO+LIRI group. LIRI included 90-min ischemia and 120-min reperfusion, while RhEpo was administered (3 kU/kg) intraperitoneally 2 h before the operation. Levels of pulmonary inflammatory responses were examined by analyzing pulmonary permeability index (PPI), oxygenation index, histology, and expressions of inflammatory cytokines. RESULTS Pretreatment with EPO significantly decreased lung W/D ratio, BALF leukocytes count and percentage, and PPI but increased oxygenation index compared with the LIRI group (P<0.05). More importantly, with EPO pretreatment there was less pathological damage compared with the vehicle group. Expressions of inflammatory cytokines (TNF-α, IL-6, and IL-1β) in the serum were significantly lower in the EPO group than in the LIRI group (P<0.05). In addition, gene expression and protein expression of TLR-4 and NF-κB were significantly inhibited with EPO pretreatment compared with the LIRI group (P<0.05). CONCLUSIONS Our study id the first to report that EPO protects lung injuries after LIRI through inhibiting the TLR4-NF-κB signaling pathway, which provides solid evidence for the use of EPO as a therapeutic agent for treating LIRI in the future.

Topics: Animals; Cytokines; Erythropoietin; Lung; Male; NF-kappa B; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction; Toll-Like Receptor 4

Congenital urinary tract obstruction is one of the most frequent malformations in fetuses or neonates, which usually causes profound impairment of renal function including reductions in both glomerular filtration rate (GFR) and tubular handling of water and solutes. Although obstruction can be released by surgical operation, the child will suffer from diuresis for sometime. It has been reported that erythropoietin (EPO) could prevent the down-regulation of aquaporin-2 (AQP2) and urinary-concentrating defects induced by renal ischemia/reperfusion (I/R) injury. However, whether EPO could promote the recovery of renal function and AQP2 expression after releasing of ureteral obstruction has not been reported yet. The purposes of the present study were to investigate the effects of EPO on renal function and AQP2 expression after release of bilateral ureteral obstruction (BUO-R) in rats. The results showed that EPO could promote interleukin (IL) 10 (IL-10) expression; inhibit tumor necrosis factor-α (TNF-α), IL-6, and inducible nitric oxide synthase (iNOS) expressions; reduce the fractional excretion of sodium (FENa) and plasma creatinine (CREA) and urea; and promote the recovery of water and salt handling and AQP2 expression in BUO-R rats, especially in the high dose of EPO-treated group rats. In conclusion, EPO could promote the recovery of renal function and AQP2 expression in BUO-R rats, which may partially associate with its anti-inflammation effect.

Topics: Animals; Aquaporin 2; Disease Models, Animal; Erythropoietin; Glomerular Filtration Rate; Humans; Kidney; Male; Protective Agents; Rats, Sprague-Dawley; Reperfusion Injury; Ureteral Obstruction

Topics: Adenine; Animals; Autophagosomes; Autophagy; Cell Line; Chemical and Drug Induced Liver Injury; Chromones; Cobalt; Disease Models, Animal; Erythropoietin; Humans; Liver; Liver Function Tests; Mice; Mice, Inbred C57BL; Morpholines; Peptide Fragments; Proto-Oncogene Proteins c-akt; Random Allocation; Reperfusion Injury; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases

The effects of erythropoietin (EPO) alone or in conjunction with ischemic preconditioning (IPC) on nitric oxide synthase as well as comparing their effects on oxidative stress and proinflammatory cytokines are studied. Rats underwent bilateral renal ischemia of 50 min followed by 24 h reperfusion. They were administered EPO (5000 iu/kg i.p.) and/or subjected to IPC and sacrificed after 24 h, then plasma and tissue samples were obtained. Treatment of either EPO or IPC and their combination attenuates oxidative stress, decreases histological damages, inhibits proinflammatory response, and up-regulates iNOS and eNOS gene expression compared to IR group. In addition, EPO+IPC and EPO treatment produced significant up-regulation in iNOS gene expression compared to IPC group. In IPC and EPO+IPC groups, more powerful effect on up-regulation of eNOS gene expression was shown compared to EPO group. Our findings suggest that treatment with EPO or IPC and their combination improve renal function and preserve tubular damage induced by IR injury. These advantageous effects were closely related to reducing oxidative stress, suppressing proinflammatory response and enhancing generation of NO. IPC was more powerful in enhancement of eNOS gene expression compared to EPO that was more effective in increasing of iNOS gene expression.

Topics: Acute Kidney Injury; Animals; Combined Modality Therapy; Erythropoietin; Ischemic Preconditioning; Male; Nitric Oxide Synthase; Rats; Rats, Wistar; Reperfusion Injury; Treatment Outcome; Up-Regulation

Salidroside is being investigated for its therapeutic potential in stroke because it is neuroprotective over an extended therapeutic window of time. In the present study, we investigated the mechanisms underlying the anti-inflammatory effects of salidroside (50 mg/kg intraperitoneally) in rats, given 1 h after reperfusion of a middle cerebral artery that had been occluded for 2 h. After 24 h, we found that salidroside increased the neuronal nuclear protein NeuN and reduced the marker of microglia and macrophages CD11b in the peri-infarct area of the brain. Salidroside also decreased IL-6, IL-1β, TNF-α, CD14, CD44, and iNOs mRNAs. At the same time, salidroside increased the ratio of phosphorylated protein kinase B (p-Akt) to total Akt. The phosphoinositide 3-kinase (PI3K) inhibitor LY294002 prevented this increase in p-Akt and reversed the inhibitory effects of salidroside on CD11b and inflammatory mediators. Salidroside also elevated the protein levels of hypoxia-inducible factor (HIF) subunits HIF1α, HIF2α, HIF3α, and of erythropoietin (EPO). The stimulatory effects of salidroside on HIFα subunits were blocked by LY294002. Moreover, YC-1, a HIF inhibitor, abolished salidroside-mediated increase of HIF1α and prevented the inhibitory effects of salidroside on CD11b and inflammatory mediators. Taken together, our results provide evidence for the first time that all three HIFα subunits and EPO can be regulated by PI3K/Akt in cerebral tissue, and that salidroside entrains this signaling pathway to induce production of HIFα subunits and EPO, one or more of which mediate the anti-inflammatory effects of salidroside after cerebral IRI.

Topics: Animals; Anti-Inflammatory Agents; Brain Ischemia; Erythropoietin; Glucosides; Hypoxia-Inducible Factor 1, alpha Subunit; Inflammation; Phenols; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Rats; Reperfusion Injury; Signal Transduction

The human brain requires uninterrupted delivery of blood-borne oxygen and nutrients to sustain its function. Focal ischemia, particularly, ischemic stroke, and global ischemia imposed by cardiac arrest disrupt the brain's fuel supply. The resultant ATP depletion initiates a complex injury cascade encompassing intracellular Ca

Topics: Animals; Brain; Brain Ischemia; Erythropoietin; Humans; Neuroprotective Agents; Oxidative Stress; Reperfusion Injury

Methylprednisolone sodium succinate (MPSS) has been suggested as a treatment for spinal cord injury (SCI), but its use has been limited due to its adverse effects. Erythropoietin (EPO) has been suggested as a promising candidate for limiting SCI in mammals. The aim of the present study was to investigate the effects of EPO in combination with MPSS on astrocytes following ischemic injury in vitro. Astrocytes were isolated from the cerebral cortex of postnatal day 3 Sprague‑Dawley rats and cultured in vitro. Astrocyte ischemic injury was induced by oxygen and glucose deprivation for 4 h, and reperfusion was simulated by subsequent culture under normoxic conditions. The effects of EPO and MPSS on the expression of aquaporin‑4 (AQP4) were investigated. Ischemic astrocytes were treated with EPO (10 U/ml), MPSS (10 µg/ml), or EPO (10 U/ml) in combination with MPSS (10 µg/ml) during reperfusion. The cell viability of astrocytes was assessed using an MTT assay. The mRNA and protein expression levels of AQP4 were determined using reverse transcription‑quantitative polymerase chain reaction and western blot analysis, respectively. The role of the protein kinase C (PKC) signaling pathway in the molecular mechanisms underlying the effects of EPO and MPSS was also investigated. The present results demonstrated that following treatment with EPO and MPSS, the mRNA expression levels of AQP4 were upregulated and cell viability was enhanced. EPO and MPSS effectively inhibited the oxygen and glucose deprivation‑mediated downregulation of AQP4 following reperfusion. In addition, the combined treatment with EPO and MPSS exhibited higher AQP4 expression levels and cell viability compared with each treatment alone. Finally, the effects of EPO and MPSS on AQP4 expression were partially reversed by pretreatment with the PKC inhibitor Ro 31‑8220. The present study indicated that EPO and MPSS had a synergistic effect on AQP4 expression following reperfusion, and suggest that they may be combined in the treatment of SCI.

Topics: Animals; Animals, Newborn; Aquaporin 4; Astrocytes; Cell Hypoxia; Cell Survival; Erythropoietin; Female; Gene Expression Regulation; Glucose; Hypothalamus; Indoles; Methylprednisolone Hemisuccinate; Neuroprotective Agents; Oxygen; Primary Cell Culture; Protein Kinase C; Protein Kinase Inhibitors; Rats; Rats, Sprague-Dawley; Reperfusion Injury; RNA, Messenger; Signal Transduction

Remote ischemic preconditioning (RIPC) is an intriguing approach which exposes a remote organ/tissue to a non-lethal transient ischemia/reperfusion (I/R) in order to potentiate the resistance of the desired organ/tissue against the next unwanted I/R. It has been suggested that RIPC exerts its effect through neuronal and hormonal pathways. The underlying mechanisms of RIPC are obscure and should be elucidated. In this study, we induced RIPC in mice using 3 cycles of 5 min ischemia alternating with 5 min reperfusion of the left renal artery. Renal failure was induced in mice by intra-peritoneal (i.p.) injection of 200 mg/kg body weight of gentamicin twice per day for 4 consecutive days. Global hippocampal ischemia reperfusion (I/R) was performed by bilateral carotid artery occlusion for 20 min followed by reperfusion for 72 h. Moreover, the retention trial of passive avoidance test was determined 72 h after global ischemia. Histopathological changes of hippocampus neurons were observed using Nissl staining to detect neuronal loss. Finally, terminal deoxynucleotidyl transferase mediated dUTP nick end-labeling (TUNEL) was performed to assess the status of apoptotic cells in the hippocampus. The results of this study suggest that renal ischemic preconditioning is a good candidate for prevention of I/R-induced hippocampal injury. However, RRPC (remote renal preconditioning) failed to exert a neuroprotective effect in mice with renal failure (RF), indicating the probable role of a humoral factor which is released from kidneys in response to ischemia. In agreement with this hypothesis, treatment of mice with rhEPO (5000 IU/kg intraperitoneal) before induction of RRPC restored the neuroprotective effects of RRPC in RF mice. Accordingly, it is plausible to expect that erythropoietin is released from kidneys to act as a mediator for RRPC-induced neuroprotective effects. Renal ischemic preconditioning prevents I/R-induced hippocampal injury. In contrast, renal failure hampers protective effects of RRPC, while exogenous administration of erythropoietin (EPO) significantly prevents the inhibiting effects of renal failure.

Topics: Animals; Avoidance Learning; Erythropoietin; Hippocampus; Ischemic Preconditioning; Kidney; Mice; Renal Insufficiency; Reperfusion Injury

This study compared the hematopoietic capacities of erythropoietin (Epo) and antioxidant drug U-74389G, based on 2 preliminary studies. The provided results on hematocrit levels augmentation were co-evaluated in a hypoxia reoxygenation protocol of an animal model.. Hematocrit levels were evaluated at the 60th reoxygenation min (for groups A, C and E) and at the 120th reoxygenation min (for groups B, D and F) in 60 rats. Groups A and B received no drugs, rats from groups C and D were administered with Epo; whereas rats from groups E and F were administered with U-74389G.. The first preliminary study of Epo non-significantly increased the hematocrit levels by 0.24%+1.38% (p-value=0.8586). The second preliminary study of U-74389G significantly raised the hematocrit levels by 3.16%+1.33% (p-value=0.0196). These 2 studies were co-evaluated since they came from the same experimental setting. The outcome of the co-evaluation was that U-74389G has approximately 12.66-fold higher hematopoietic potency than Epo (p-value=0.0000).. The anti-oxidant capacities of U-74389G provide satisfactory acute hematopoietic properties; presenting approximately 12.66-fold hematocrit level rise than epo (p-value=0.0000).

Topics: Animals; Antioxidants; Disease Models, Animal; Erythropoietin; Female; Hematocrit; Hematopoiesis; Humans; Hypoxia; Male; Pregnatrienes; Rats, Wistar; Reperfusion Injury

Acute ischemia-reperfusion (I/R) injury is the most common causes of acute renal failure in daily clinical\ practice. It has been recognized that endothelial cell dysfunction and microvascular injury as the pathophysiological changes during I/R injury. Protective effects of erythropoietin (EPO) have been demonstrated in various experimental models of I/R induced injury. Therefore, the aim of the present study was to investigate whether EPO administration has renoprotective effect against acute renal failure I/R injury in rats by promotion of endothelial progenitor cells (EPCs) mobilization and\ neovascularization.. Male Sprague-Dawley rats were pretreated with EPO (1,000 IU/kg/day, ip); or the placebo for 3 days before the induction of I/R procedure. On day 4, the bilateral renal occlusion for 30 min operations to produce renal I/R injury or treatment with EPO 30 min before the initiation of I/R were done. At the end of the reperfusion period at day 1 day 2 and day 4, blood and renal tissues were collected to investigate renal function and pathohistological examination. The expression levels of CAV-1 and CD34 were determined for circulating of EPCs in blood, while CD34, CAV-1 and VEGFR-2 were investigated for mobilized EPCs in kidney, using real time PCR. The expression level of VEGF was also examined to indicate the angiogenesis in kidney using real time PCR and western blotting.. In the I/R group, the significantly increased values of serum urea and creatinine were found on Day 1 after ischemia, as compared to sham group. The development of tubular epithelial cell necrosis, peritubular capillary congestion and mild interstitial infiltration has been observed in this group. Administration of EPO in I/R rat was significantly improved renal function and significantly less the tubular damage. The treatment with EPO significantly increased in expression levels of CD34 and CAV-1 in blood, and also CAV-1, VEGFR-2 and VEGF in kidney tissue in this group, as compared to the I/R group.. These results suggest that treatment with EPO protects the kidney from ischemic acute renal injury via\ increasing the mobilization and recruitment of EPCs, resulting in the induction of expression of VEGF that might play an important role in the repair response.

Topics: Acute Kidney Injury; Animals; Blood Urea Nitrogen; Creatinine; Disease Models, Animal; Erythropoietin; Ischemia; Kidney; Male; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2

This study examined effects and functional outcome of recombinant human erythropoietin (rhEPO) and carbamylated erythropoietin fusion protein (cEPO-FC) preconditioning in a rabbit model for spinal cord ischemia and resulting paraplegia. This model was chosen because only a small surgical effect is needed to cause paraplegia in rabbits, which facilitates postoperative observation of animals.. Anesthetized but spontaneously breathing New Zealand White rabbits randomly received cEPO-FC (50 μg/kg; n = 8), rhEPO (5000 IU/kg; n = 10), or vehicle (control; n = 10) 30 minutes before and after infrarenal aortic clamping. Ideal clamping time of 22 minutes was identified from preceding clamping tests (15-25 minutes). Postoperative observation time was 96 hours. Spinal cord function was assessed by neurologic evaluation of hind limb motor function every 12 hours using a modified Tarlov score. Spinal cord tissue damage was evaluated after 96 hours using hematoxylin and eosin, elastica van Gieson, Nissl, Masson-Goldner, and hemosiderin staining. Plasma levels of cell senescence markers stathmin, chitinase 1/3, elongation factor 1-α were determined.. Rabbits that received rhEPO showed significant improvement of spontaneous lower limb movements until 36 hours of reperfusion and improved histologic scores upon examination of the lumbar spinal cord compared with the control group. In contrast, cEPO-FC treatment showed comparable outcome to the control group concerning movements of the lower limbs and histology. Senescence markers were elevated in the control group, but not in the treatment groups, except for chitinase 3 in the rhEPO group. Only stathmin showed no significant effect. Markers for senescence might increase after acute ischemic injury. Attenuation of senescence markers might not come alone from improvement of the spinal cord.. Preconditioning with rhEPO attenuates ischemia/reperfusion injury of the spinal cord, whereas the carbamylated derivative (cEPO-FC) showed no positive effect on spinal cord function.

Topics: Animals; Biomarkers; Cellular Senescence; Chitinases; Disease Models, Animal; Erythropoietin; Male; Motor Activity; Neurologic Examination; Neuroprotective Agents; Paraplegia; Peptide Elongation Factor 1; Rabbits; Recombinant Proteins; Reperfusion Injury; Spinal Cord; Spinal Cord Ischemia; Stathmin; Time Factors

Neurologic sequelae remain a common and destructive problem in patients with acute kidney injury. The objective of this study was to evaluate the possible neuroprotective effect of erythropoietin (EPO) on motor impairments following bilateral renal ischemia (BRI) in two time points after reperfusion: short term (24 h) and long term (1 week). Male Wistar rats underwent BRI or sham surgery. EPO or saline administration was performed 30 min before surgery (1000 U/kg, i.p.). Explorative behaviors and motor function of the rats were evaluated by open field, rotarod, and wire grip tests. Plasma concentrations of blood urea nitrogen (BUN) and creatinine (Cr) were significantly enhanced in BRI rats 24 h after reperfusion. BRI group had only an increased level of BUN but not Cr 1 week after reperfusion. Impairment of balance function by BRI was not reversed by EPO 24 h after reperfusion, but counteracted 7 days after renal ischemia. Muscle strength had no significant differences between the groups. BRI group had a decrease in locomotor activity, and EPO could not reverse this reduction in both time points of the experiment. Although EPO could not be offered as a potential neuroprotective agent in the treatment of motor dysfunctions induced by BRI, it could be effective against balance dysfunction 1 week after renal ischemia.

Topics: Acute Kidney Injury; Animals; Blood Urea Nitrogen; Creatinine; Disease Models, Animal; Erythropoietin; Locomotion; Male; Motor Disorders; Muscle Strength; Neuroprotective Agents; Rats; Rats, Wistar; Reperfusion; Reperfusion Injury

Renal ischemia-reperfusion (IR) contributes to the development of acute renal failure (ARF). Oxygen free radicals are considered to be principal components involved in the pathophysiological tissue alterations observed during renal IR. The purpose of this study was to investigate the combination effect of melatonin (MEL) and erythropoietin (EPO), which are a potent antioxidant and anti-apoptotic agents, in IR-induced renal injury in rats. Wistar Albino rats were unilaterally nephrectomized and subjected to 45 min of renal pedicle occlusion followed by 24 h reperfusion. MEL (10 mg/kg, i.p) and EPO (5000 U/kg, i.p) were administered prior to ischemia. After 24 h reperfusion, following decapitation, blood samples were collected for the determination of superoxide dismutase (SOD), glutathione peroxidase (GPx), and malondialdehyde (MDA) levels. Also, renal samples were taken for histological evaluation and apoptosis assay. Ischemia-reperfusion increased SOD, GPx, MDA levels, and TUNEL positive cells. Histopathological findings of the IR group confirmed that there was renal impairment in the tubular epithelium. Treatment with EPO and MEL decreased SOD, GPx, and MDA levels, histopathological changes, and TUNEL positive cells. These results indicated that the combination of MEL and EPO could not exert more nephroprotective and anti-apoptotic effects than MEL treatment in renal ischemia-reperfusion injury.

Topics: Acute Kidney Injury; Animals; Antioxidants; Apoptosis; Epoetin Alfa; Erythropoietin; Glutathione Peroxidase; Male; Malondialdehyde; Melatonin; Rats; Rats, Wistar; Reperfusion Injury; Superoxide Dismutase

Intra-artery infusion of recombinant human erythropoietin (rhEPO) has recently been reported to confer neuroprotection against cerebral ischemia-reperfusion injury in animal models; however, the molecular mechanisms are still under investigation. The present study focused on the specific mechanism involved in blood-brain barrier (BBB) disruption.. Thirty-six male and nine female Sprague Dawley rats were subjected to middle cerebral artery (MCA) occlusion to induce focal cerebral ischemia, and administrated rhEPO at a dose of 800 U/kg through MCA infusion at the beginning of reperfusion. Neurobehavioral deficits, brain edema, and infarct volume were evaluated after 2 h of ischemia and 24 h of reperfusion. BBB permeability was assessed by quantifying the extravasation of Evans blue (EB) dye. The expression of tight junction proteins and matrix metalloproteinases (MMPs) (Claudin-5, Occludin, MMP-2, and MMP-9) in microvessels were detected by immunofluorescence and western blot. The activities of MMPs in the cerebral microvessels were determined by gelatin zymography.. Treatment with rhEPO through the MCA strongly alleviated infarct volume, brain edema, and improved neurobehavioral outcomes in male and female rats. In addition, rhEPO remarkably suppressed the EB extravasation induced by brain ischemia. Furthermore, rhEPO prevented degradation of Claudin-5 and Occludin, and reduced the expression and activity of MMP-2 and MMP-9 in isolated brain microvessels.. Treatment with rhEPO through MCA infusion prevented brain edema formation and infarction through inhibition of MMP-mediated BBB disruption in acute ischemic stroke.

Topics: Animals; Blood-Brain Barrier; Brain Edema; Brain Ischemia; Cerebral Infarction; Erythropoietin; Female; Humans; Infarction, Middle Cerebral Artery; Infusions, Intra-Arterial; Male; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury

Paraplegia remains a devastating complication of complex aortic surgery. Erythropoietin (EPO) has been shown to prevent paraplegia after ischemia reperfusion, but the protective mechanism remains poorly described in the spinal cord. We hypothesized that EPO induces the CREB (cAMP [adenosine 3'5' cyclic monophosphate] response element-binding protein) pathway and neurotrophin production in the murine spinal cord, attenuating functional and cellular injury.. Adult male mice were subjected to 4 minutes of spinal cord ischemia via an aortic and left subclavian cross-clamp. Experimental groups included EPO treatment 4 hours before incision (n = 7), ischemic control (n = 7), and shams (n = 4). Hind-limb function was assessed using the Basso motor score for 48 hours after reperfusion. Spinal cords were harvested and analyzed for neuronal viability using histology and staining with a fluorescein derivative. Expression of phosphorylated (p)AKT (a serine/threonine-specific kinase), pCREB, B-cell lymphoma 2, and brain-derived neurotrophic factor were determined using immunoblotting.. By 36 hours of reperfusion, EPO significantly preserved hind-limb function after ischemia-reperfusion injury (P < .01). Histology demonstrated preserved cytoarchitecture in the EPO treatment group. Cords treated with EPO expressed significant increases in pAKT (P = .021) and pCREB (P = .038). Treatment with EPO induced expression of both of the neurotrophins, B-cell lymphoma 2, and brain-derived neurotrophic factor, beginning at 12 hours.. Erythropoietin-mediated induction of the CREB pathway and production of neurotrophins is associated with improved neurologic function and increased neuronal viability following spinal cord ischemia reperfusion. Further elucidation of EPO-derived neuroprotection will allow for expansion of adjunct mechanisms for spinal cord protection in high-risk thoracoabdominal aortic intervention.

Topics: Animals; Brain-Derived Neurotrophic Factor; CREB-Binding Protein; Disease Models, Animal; Erythropoietin; Male; Mice, Inbred C57BL; Motor Activity; Paraplegia; Phosphorylation; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-bcl-2; Reperfusion Injury; Signal Transduction; Spinal Cord; Spinal Cord Ischemia; Time Factors

Renal ischaemia-reperfusion (I/R) injury, a primary cause of acute renal failure, can induce high morbidity and mortality. This study aimed to explore the effect of erythropoietin on renal I/R injury and its underlying mechanism.. Fifty male Sprague-Dawley rats were randomly allocated to three groups (n = 10): the sham group, the renal ischaemia-reperfusion-saline (IRI) group, and the IRI+-Erythropoietin (EPO) group. Erythropoietin (250, 500, 1000 U/kg) was intraperitoneally injected 30 min before inducing I/R. Renal I/R injury were induced by clamping the left renal artery for 30 min followed by reperfusion, along with a contralateral nephrectomy. Renal function and histological damage were determined after 24 h reperfusion. The expression of pro-inflammatory cytokines interleukin-6 (IL-6), interleukin-1 β (IL-1β), and tumour necrosis factor-α (TNF-α) in the serum and renal tissue were evaluated by enzyme linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR), respectively. Further, the effects of erythropoietin on PI3K/Akt signalling, erythropoietin receptor (EPOR) and nuclear factor (NF)-κB activation were measured by Western blotting.. Erythropoietin pretreatment can significantly decrease the level of renal dysfunction in a dose-dependent manner, attenuated the renal histological changes, the expression of TNF-α, IL-1β, and IL-6, the levels of reactive oxygen species (ROS) production and NF-κB p65 phosphorylation in renal tissue upon IRI. Moreover, erythropoietin pretreatment could further activate the PI3K/Akt signalling and induced EPOR activity.. Erythropoietin pretreatment could attenuate renal I/R injury by suppressing inflammation, which was associated with activating PI3K/Akt signalling though EPOR activation. Our findings suggest that erythropoietin may be a novel practical strategy to prevent renal I/R injury.

Topics: Acute Kidney Injury; Animals; Anti-Inflammatory Agents; Biomarkers; Cytoprotection; Disease Models, Animal; Enzyme Activation; Erythropoietin; Inflammation Mediators; Interleukin-1beta; Interleukin-6; Kidney; Male; Phosphatidylinositol 3-Kinase; Phosphorylation; Proto-Oncogene Proteins c-akt; Rats, Sprague-Dawley; Reactive Oxygen Species; Receptors, Erythropoietin; Reperfusion Injury; Signal Transduction; Transcription Factor RelA; Tumor Necrosis Factor-alpha

Topics: Animals; CREB-Binding Protein; Erythropoietin; Male; Paraplegia; Reperfusion Injury; Signal Transduction; Spinal Cord; Spinal Cord Ischemia

Local infusion of low dose erythropoietin (EPO) alleviates cerebral ischemia and reperfusion (I/R) injury in rats; however, the underlying molecular mechanisms are still unclear. The present study investigated the effect of low dose EPO treatment on I/R-induced endoplasmic reticulum (ER) stress in brain tissue and isolated microvessels in rodents. Sprague-Dawley rats were subjected to 2 h ischemia/24 h reperfusion by middle cerebral artery (MCA) occlusion, then administered fluorescein isothiocyanate-labeled EPO via MCA infusion (MCAI) or subcutaneous injection (SI) to compare the efficiency of two modes of delivery. Neurobehavioral deficits and infarct volume, and the expression of ER stress-associated proteins and apoptosis in brain tissue or isolated microvessels, as well as the transcriptional activity of 16 factors involved in ER stress and the unfolded protein response in brain tissue was asscessed. A higher EPO level in cerebrospinal fluid and brain tissue was observed in rats treated with EPO by MCAI (800 IU/kg) than by SI (5000 IU/kg). Moreover, neurobehavioral deficits and infarct volume were reduced in rats treated with EPO by MCAI and salubrinal. EPO suppressed the expression of ER stress signals glucose-regulated protein 78, activating transcription factor (ATF) 6α, and CCAAT enhancer-binding protein homologous protein (CHOP), as well as that of the pro-apoptotic protein caspase-3 in brain microvessels, and decreased the number of CHOP-positive, apoptotic neurons. EPO treatment also reduced the transcriptional activities of CHOP, forkhead box protein O1, and ATF4. These results provide evidence that low dose EPO treatment via MCAI provides neuroprotection following acute ischemic stroke by inhibiting the ER stress response.

Topics: Animals; Apoptosis Regulatory Proteins; Brain Ischemia; Capillaries; Cerebrovascular Circulation; Endoplasmic Reticulum Stress; Epoetin Alfa; Erythropoietin; Heat-Shock Proteins; Infarction, Middle Cerebral Artery; Infusions, Intra-Arterial; Male; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury

Understanding the mechanisms of protecting the kidneys from injury is of great importance because there are no effective therapies that promote repair and the kidneys frequently do not repair adequately. Evidence has shown that erythropoietin (EPO) has a vital renoprotective role, independent of its erythropoietic effect. However, whether EPO can contribute to kidney repair after injury and the potential mechanisms are not fully understood.. To investigate the renoprotective mechanism of EPO, a kidney ischemia/reperfusion injury (IRI) model was induced in adult male Sprague-Dawley rats. The rats were subsequently randomly treated with EPO or a vehicle 6 hours after the kidney IRI. The rats were sacrificed on Day 3, Day 5, and Day 7 post kidney IRI. Renal function and histological alterations were examined. Renal interstitial macrophage infiltration, cell proliferation, apoptosis, and angiogenesis were evaluated by immunostaining. Furthermore, the effects of EPO on the Wnt/β-catenin pathway and IRI-related micro-RNAs were investigated.. The administration of EPO significantly improved renal function and reduced tubular injury. Furthermore, EPO treatment significantly prevented tubular cell apoptosis and promoted cell proliferation after IRI. Erythropoietin significantly suppressed macrophage infiltration, compared to the vehicle. In addition, treatment with EPO markedly prevented the loss of microvasculature. We have also demonstrated that, compared to the vehicle, EPO administration enhanced the expression of Wnt7b and β-catenin, and downregulated miR-21, -214, -210, and -199a.. Erythropoietin protects the kidneys against IRI by attenuating injury of the renal microvasculature and tubule epithelial cells, by promoting Wnt/β-catenin pathway activation, and by regulating miRNA expression.

Topics: Acute Kidney Injury; Animals; Apoptosis; Disease Models, Animal; Erythropoietin; Kidney; Male; MicroRNAs; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Wnt Signaling Pathway

The inflammatory response plays an important role in liver dysfunction after hepatic ischemia/reperfusion (I/R), which is tightly regulated by the Toll-like receptor 2 (TLR2)/nuclear factor (NF)-κB pathway; suppression of TLR2/NF-κB signaling has therefore become a promising target for anti-inflammatory treatment in hepatic I/R injury. Erythropoietin (EPO) is a glycoprotein cytokine produced primarily by the kidney that has anti-inflammatory activities. The purpose of the present study was to investigate the effect of EPO preconditioning, if any, against hepatic I/R injury in rats and its underlying mechanisms.. Male Sprague-Dawley rats were subjected to partial (70%) hepatic ischemia for 45 minutes after pretreatment with either saline or EPO followed by 24-hour reperfusion. Hepatic injury was evaluated according to biochemical and histopathologic examinations. The expression of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) were measured by using enzyme-linked immunosorbent assay and real-time polymerase chain reaction. The expression of nuclear translocation and phosphorylation of NF-κB p65, EPOR receptor (EPOR), p-EPOR, p-IκB-α, IκB-α, and TLR2 were determined by using Western blot analysis.. EPO treatment significantly improved hepatic function and histology, as indicated by reduced transaminase levels and pathologic changes. The expression of TNF-α, IL-1β, IL-6, p-IκB-α, and TLR2 was significantly decreased with up-regulation of p-EPOR by EPO. Moreover, EPO pretreatment also reduced I/R-induced the phosphorylation and nuclear translocation of NF-κB p65 subunits in liver tissue, but EPO had no influence on the expression of p65 and IκB-α.. These results suggest that EPO pretreatment ameliorates hepatic I/R injury, which is involved in suppressing TLR2/NF-κB-mediated inflammation.

Topics: Animals; Enzyme-Linked Immunosorbent Assay; Erythropoietin; I-kappa B Proteins; Inflammation; Interleukin-1beta; Interleukin-6; Ischemic Preconditioning; Liver; Male; NF-KappaB Inhibitor alpha; Peptide Fragments; Phosphorylation; Rats; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; Receptors, Erythropoietin; Reperfusion Injury; Signal Transduction; Toll-Like Receptor 2; Transcription Factor RelA; Tumor Necrosis Factor-alpha; Up-Regulation

To explore the mechanism whereby stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) jointly mobilize bone marrow stem cells (BMSCs) and promote kidney repair, male Sprague-Dawley rats were randomly assigned into 4 groups. In the treatment control group, rats were administered SCF (200 μg·kg(-1)·day(-1)) and G-CSF (50 μg·kg-1·day-1) for 5 days. In the treatment group, RIRI models were established, and 6 h later, SCF (200 μg·kg(-1)·day(-1)) and G-CSF (50 μg·kg(-1)·day(-1)) were administered for 5 days. In the model and treatment groups, tubular epithelial cell degeneration and necrosis were noticed, but the extent of repair in the treatment group was significantly better than in the model group. Five days after the operation, renal tissue CD34+ cells significantly increased in the model and treatment groups compared with the control and treatment control groups. HIF-1α, VEGF, and EPO expression in treatment groups increased significantly compared with the other groups. HIF- 1α, VEGF, EPO expression in the treatment control group increased significantly compared with the control group. Joint use of SCF and G-CSF increased the number of BMSCs in damaged kidney tissue and reduced the degree of renal tissue damage. BMSCs promote increased HIF-1α expression in renal tissue. Increased kidney tissue HIF- 1α and its target gene products VEGF and EPO expression possibly induce SCF and G-CSF to promote acute tubular necrosis repair.

Topics: Animals; Bone Marrow Cells; Erythropoietin; Granulocyte Colony-Stimulating Factor; Hematopoietic Stem Cells; Hypoxia-Inducible Factor 1, alpha Subunit; Kidney; Male; Random Allocation; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Stem Cell Factor; Vascular Endothelial Growth Factor A

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.. 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.. 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.. 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/normal conditions.

Topics: Apoptosis; Cell Line; Cell Proliferation; Diazoxide; Erythropoietin; Gene Expression Regulation; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; KATP Channels; Kidney; Kidney Diseases; Kidney Tubules; Reperfusion Injury

Ischemia reperfusion injury (IRI) is a primary concern in liver transplantation, especially when steatosis is present. Acetazolamide (AZ), a specific carbonic anhydrase (CA) inhibitor, has been suggested to protect against hypoxia. Here, we hypothesized that AZ administration could be efficient to protect fatty livers against cold IRI. Obese Zucker rat livers were preserved in Institut Georges Lopez-1 storage solution for 24 hours at 4°C and ex vivo perfused for 2 hours at 37°C. Alternatively, rats were also treated with intravenous injection of AZ (30 mg/kg) before liver recovery. Liver injury, hepatic function, and vascular resistance were determined. CA II protein levels and CA hydratase activity were assessed as well as other parameters involved in IRI (endothelial nitric oxide synthase, mitogen activated protein kinase family, hypoxic inducible factor 1 alpha, and erythropoietin). We demonstrated that AZ administration efficiently protects the steatotic liver against cold IRI. AZ protection was associated with better function, decreased vascular resistance, and activation of endothelial nitric oxide synthase. This was consistent with an effective mitogen activated protein kinase inactivation. Finally, no effect on the hypoxic inductible factor 1 alpha/erythropoietin pathway was observed. The present study demonstrated that AZ administration is a suitable pharmacological strategy for preserving fatty liver grafts against cold IRI.

Topics: Acetazolamide; Animals; Carbonic Anhydrase II; Carbonic Anhydrase Inhibitors; Cold Temperature; Enzyme Activation; Erythropoietin; Extracellular Signal-Regulated MAP Kinases; Fatty Liver; Hypoxia-Inducible Factor 1, alpha Subunit; JNK Mitogen-Activated Protein Kinases; Liver; Liver Transplantation; Male; Nitric Oxide Synthase Type III; Organ Preservation; Phosphorylation; Rats, Zucker; Reperfusion Injury; Vascular Resistance

Ischemia/reperfusion injury (IRI) has lzong been an area of concern and focus of investigations. Erythropoietin (EPO) exhibits multiple protective effects, and selenium is an antioxidant trace element in the body, however, there have been no reports concerning the effects of EPO combined with sodium selenite on IRI. In the present study, a mouse model of renal IRI (RIRI) was pre‑treated with EPO and sodium selenite to determine the most appropriate combination ratio of the two for further investigation. The results revealed that EPO and sodium selenite had synergistic protective effects in RIRI. EPO was identified as the predominant treatment component, with sodium selenite serving as an adjuvant, and combination treatment was markedly more effective, compared with treatment with either drug alone. The optimal ratio of treatment was 10:1 (10 IU EPO: 1 µg sodium selenite). The results indicated that RIRI markedly induced renal injury, as evidenced by elevated levels of blood urea nitrogen (BUN), as well as higher pathological scores, based on hematoxylin and eosin staining. Pre‑treatment with EPO and sodium selenite significantly decreased serum expression levels of BUN and malonaldehyde, and increased the expression levels of superoxide dismutase, glutathione peroxidase and nitric oxide (NO), compared with the model group. Furthermore, co‑treatment with EPO and sodium selenite upregulated the protein expression levels of phosphatidylinositol‑3 kinase (PI3K) in renal tissue samples. Together, the results suggested that co‑administration of EPO and sodium selenite effectively ameliorates IRI‑induced renal injury by reducing oxidative stress and activating the PI3K/NO signaling pathway.

Topics: Animals; Blood Urea Nitrogen; Drug Synergism; Drug Therapy, Combination; Erythropoietin; Glutathione Peroxidase; Immunohistochemistry; Kidney; Male; Mice; Mice, Inbred Strains; Nitric Oxide; Oxidative Stress; Phosphatidylinositol 3-Kinase; Protective Agents; Reperfusion Injury; Sodium Selenite; Superoxide Dismutase

We developed a novel, erythropoietin-derived, non-erythropoiesis, cyclic helix B peptide (CHBP) that displays potent renoprotection against acute kidney injury (AKI). To determine the mechanism of CHBP-mediated protection, we investigated the proteomic profile of mice treated with CHBP in a kidney ischemia-reperfusion (IR) injury model. The isobaric tags for relative and absolute quantitation (iTRAQ)-labeled samples were analyzed using a QSTAR XL LC/MS system. In total, 38 differentially expressed proteins (DEPs) were shared by all experimental groups, while 3 DEPs were detected specifically in the IR + CHBP group. Eight significant pathways were identified, and oxidative phosphorylation was shown to be the most important pathway in CHBP-mediated renoprotection. The significant DEPs in the oxidative phosphorylation pathway elicited by CHBP are NADH-ubiquinone oxidoreductase Fe-S protein 6 (NDUFS6), alpha-aminoadipic semialdehyde synthase (AASS) and ATP-binding cassette sub-family D member 3 (ABCD3). The DEPs mentioned above were verified by RT-qPCR and immunostaining in mouse kidneys. We tested 6 DEPs in human biopsy samples from kidney transplant recipients. The trend of differential expression was consistent with that in the murine model. In conclusion, this study helps to elucidate the pharmacological mechanisms of CHBP before clinical translation.

Topics: Acute Kidney Injury; Animals; Enkephalins; Erythropoietin; Kidney; Male; Mice; Mice, Inbred BALB C; NAD; Oxidative Phosphorylation; Protein Precursors; Proteome; Proteomics; Reperfusion Injury; Signal Transduction

Topics: Animals; Erythropoietin; Kidney Diseases; Lung Diseases; Male; Oxidative Stress; Reperfusion Injury

Tissue damage in testicular torsion/detorsion is caused not only by the ischemia, but also by the ischemia/reperfusion injury after detorsion. Erythropoietin and sildenafil are considered to protect against ischemia/reperfusion injury. Here, we studied and compared their actions in testicular torsion/detorsion in adult rats.. Twenty-two adult male Wistar Albino rats were divided into four groups. Rats in group A (n = 5) were sham operated. Rats in group B (n = 5), group C (n = 6) and group D (n = 6) underwent torsion of the right testis and detorsion after 90 min. No pharmaceutical intervention was performed in group B. Erythropoietin (1,000 IU/kg) and sildenafil (0.7 mg/kg) were injected intraperitoneally in groups C and D, respectively, after 60 min of torsion. All animals were killed 24 h after detorsion, and their right testis was extracted, placed into 10 % formalin solution and sent for histopathological examination. The histological changes in the testes were scored according to the four-point grading system proposed by Cosentino et al.. All rats in group A had normal testicular architecture (grade 1). The untreated group B had a mean grade of 3.81 (range 3.65-4). The treated groups C (mean grade 3.24; range 3.05-3.45) and D (2.69, range 2.4-2.9) presented statistically significant better results (lower grades) compared with the untreated group B. Group D had significantly better results (lower grades) than group C.. The intraperitoneal injection of erythropoietin and sildenafil protects against ischemia/reperfusion injury after testicular torsion and detorsion. Sildenafil may have a stronger action than erythropoietin at the doses used in this study.

Topics: Animals; Erythropoietin; Injections, Intraperitoneal; Male; Phosphodiesterase 5 Inhibitors; Piperazines; Purines; Rats, Wistar; Reperfusion Injury; Sildenafil Citrate; Spermatic Cord Torsion; Sulfones; Testis

Treatment of renal ischemia-reperfusion (IR) injury with recombinant human erythropoietin (rhEPO) reduces acute kidney injury and improves function. We aimed to investigate whether progression to chronic kidney disease associated with acute injury was also reduced by rhEPO treatment, using in vivo and in vitro models. Rats were subjected to bilateral 40-min renal ischemia, and kidneys were studied at 4, 7, and 28 days postreperfusion for renal function, tubular injury and repair, inflammation, and fibrosis. Acute injury was modulated using rhEPO (1,000 or 5,000 IU/kg, intraperitoneally) at the time of reperfusion. Renal tubular epithelial cells or fibroblasts in culture were subjected to hypoxia or oxidative stress, with or without rhEPO (200 IU/ml), and fibrogenesis was studied. The results of the in vivo model confirmed functional and structural improvement with rhEPO at 4 days post-IR (P < 0.05). At 7 days post-IR, fibrosis and myofibroblast stimulation were increased with IR with and without rhEPO (P < 0.01). However, at 28 days post-IR, renal fibrosis and myofibroblast numbers were significantly greater with IR plus rhEPO (P < 0.01) compared with IR only. Mechanistically, rhEPO stimulated profibrotic transforming growth factor-β, oxidative stress (marker 8-hydroxy-deoxyguanosine), and phosphorylation of the signal transduction protein extracellular signal-regulated kinase. In vitro, rhEPO protected tubular epithelium from apoptosis but stimulated epithelial-to-mesenchymal transition and also protected and activated fibroblasts, particularly with oxidative stress. In summary, although rhEPO was protective of renal function and structure in acute kidney injury, the supraphysiological dose needed for renoprotection contributed to fibrogenesis and stimulated chronic kidney disease in the long term.

Topics: Acute Kidney Injury; Animals; Apoptosis; Cells, Cultured; Disease Progression; Epoetin Alfa; Erythropoietin; Extracellular Signal-Regulated MAP Kinases; Fibrosis; Humans; Kidney; Kidney Diseases; Male; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Treatment Outcome

Renal ischemia reperfusion (IR) is an important cause of renal dysfunction. It contributes to the development of acute renal failure. Oxidative damage from reactive oxygen species is considered to be the principal component involved in the pathophysiological tissue alterations observed during IR. The purpose of this study was to evaluate the effect of a combined treatment with erythropoietin (EPO) plus melatonin (MEL), which are known anti-inflammatory and antioxidant agents, in IR-induced renal injury in rats.. Wistar Albino rats were unilaterally nephrectomized and subjected to 45 min of renal pedicle occlusion followed by 24 h of reperfusion. MEL (10 mg/kg, i.p) and EPO (5000 U/kg, i.p) were administered prior to ischemia. After 24 h of reperfusion, blood samples were collected for the determination of superoxide dismutase (SOD), glutathione peroxidase (GPx), plasma levels of total antioxidant capacity (TAC), and malondialdehyde (MDA) and serum urea level. Also, renal samples were taken for histological evaluation.. Ischemia reperfusion significantly increased urea, blood SOD, and GPx levels. Histological findings of the IR group indicated that there was increase in tubular and glomerular hyaline cast, thickening of Bowman capsule basement membrane, and renal impairment in the glomerular epithelium. Treatment with EPO and MEL significantly decreased blood SOD, GPx, and urea levels and increased TAC level. In the EPO + MEL group, while the histopathological changes were lower than those in EPO group, they were the same as MEL group.. EPO and MEL combination treatment exerted more nephroprotective effects than EPO treatment and nearly had protective effects similar to MEL treatment.

Topics: Acute Kidney Injury; Animals; Anti-Inflammatory Agents; Antioxidants; Disease Models, Animal; Drug Therapy, Combination; Erythropoietin; Glutathione Peroxidase; Kidney; Male; Malondialdehyde; Melatonin; Models, Animal; Rats; Rats, Wistar; Reactive Oxygen Species; Reperfusion Injury; Superoxide Dismutase

Acute kidney injury is common, serious with no specific treatment. Ischemia-reperfusion is a common cause of acute kidney injury (AKI). Clinical trials suggest that preoperative erythropoietin (EPO) or remote ischemic preconditioning may have a renoprotective effect. Using a porcine model of warm ischemia-reperfusion-induced AKI (40-min bilateral cross-clamping of renal arteries, 48-h reperfusion), we examined the renoprotective efficacy of EPO (1,000 iu/kg iv.) or remote ischemic preconditioning (3 cycles, 5-min inflation/deflation to 200 mmHg of a hindlimb sphygmomanometer cuff). Ischemia-reperfusion induced significant kidney injury at 24 and 48 h (χ(2), 1 degree of freedom, >10 for 6/7 histopathological features). At 2 h, a panel of biomarkers including plasma creatinine, neutrophil gelatinase-associated lipocalin, and IL-1β, and urinary albumin:creatinine could be used to predict histopathological injury. Ischemia-reperfusion increased cell proliferation and apoptosis in the renal cortex but, for pretreated groups, the apoptotic cells were predominantly intratubular rather than interstitial. At 48-h reperfusion, plasma IL-1β and the number of subcapsular cells in G2-M arrest were reduced after preoperative EPO, but not after remote ischemic preconditioning. These data suggest an intrarenal mechanism acting within cortical cells that may underpin a renoprotective function for preoperative EPO and, to a limited extent, remote ischemic preconditioning. Despite equivocal longer-term outcomes in clinical studies investigating EPO as a renoprotective agent in AKI, optimal clinical dosing and administration have not been established. Our data suggest further clinical studies on the potential renoprotective effect of EPO and remote ischemic preconditioning are justified.

Topics: Acute Kidney Injury; Animals; Creatinine; Disease Models, Animal; Epoetin Alfa; Erythropoietin; Female; Hindlimb; In Situ Nick-End Labeling; Ischemic Preconditioning; Preoperative Care; Recombinant Proteins; Reperfusion Injury; Swine

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.

Topics: Animals; Behavior, Animal; Blood-Brain Barrier; Brain Ischemia; Drug Delivery Systems; Erythropoietin; Gait; Infarction, Middle Cerebral Artery; Male; Microbubbles; Neurons; Neuroprotective Agents; Rats; Rats, Wistar; Reperfusion Injury; Sound

Topics: Animals; Erythropoietin; Kidney Diseases; Lung Diseases; Male; Oxidative Stress; Reperfusion Injury

Hepatic ischemia-reperfusion (IR) results in progressive injury; initiated by oxidative stress during ischemia and compounded by cytokine-mediated inflammation during reperfusion. Recovery requires strict regulation of these events. Recombinant human erythropoietin (rhEPO) is thought to mitigate hepatocellular IR injury by altering the nonparenchymal liver microenvironment. This study sought to identify additional mechanisms whereby rhEPO is protective after liver IR injury. Mice were treated with rhEPO (4 units/g s.c.) at the onset of partial liver ischemia and assessed for transaminase and histologic injury at intervals after reperfusion. Induction of cytokines, activation of signal transducers and activators of transcription (STATs), suppressors of cytokine signaling (Socs1, Socs3, Cis), caspase-3 activation, and heme oxygenase-1 (HO-1) expression were assessed in postischemic liver. Effects of rhEPO stimulation were further characterized in whole-liver lysates from mice undergoing rhEPO injection alone and in cultured AML-12 hepatocytes. Recombinant human erythropoietin treatment at the onset of severe (90 min) hepatic IR confirmed commensurate biochemical and histological protection without affecting tissue cytokine levels. Although Socs3 and STAT5 activation were induced in normal liver after in vivo rhEPO injection, this treatment did not augment expression beyond that seen with IR alone, and neither was induced in cultured hepatocytes treated with rhEPO. Recombinant human erythropoietin inhibited caspase-3 activation in nonparenchymal cells, whereas hepatocellular HO-1 was rapidly induced both in vivo and in vitro with rhEPO treatment. These data suggest HO-1 as a potent mechanism of rhEPO-mediated protection after liver IR, which involves both direct hepatocellular and nonparenchymal mechanisms.

Topics: Animals; Apoptosis; Cells, Cultured; Cytokines; Erythropoietin; Heme Oxygenase-1; Hepatocytes; Inflammation Mediators; Liver; Male; Membrane Proteins; Mice, Inbred C57BL; Oxidative Stress; Recombinant Proteins; Reperfusion Injury; Signal Transduction; STAT Transcription Factors; Suppressor of Cytokine Signaling Proteins

Acute renal tubular injury is a serious complication in the postoperative period, which is associated with high mortality and increased ICU stay. We aimed to demonstrate the protective effect of rhEPO against acute tubular injury induced by ischemia-reperfusion and to explore the mechanism of canonical transient receptor potential channel-6.. Randomized laboratory animal study.. Animal research laboratory.. Male Sprague-Dawley rats were randomly divided into three groups: the sham group, the control group, and the rhEPO group. Experimental acute tubular injury was established in rats by bilateral renal arterial occlusion for 30 minutes followed by reperfusion.. Blood samples were obtained for cystatin-C and neutrophil gelatinase-associated lipocalin measurements by enzyme-linked immunosorbance assays. Seventy-two hours after reperfusion, urine samples were collected for osmolality and fractional excretion of sodium (%) assays on a chemistry analyzer. Kidneys were harvested at 24, 48, and 72 hours after reperfusion. Transient receptor potential channel-6, aquaporin-2, and Na,K-ATPase expression in collecting ducts were studied by immunofluorescence and Western blot. Coimmunoprecipitations were also performed to identify the possible signalplex relation between transient receptor potential channel-6 and aquaporin-2 or Na,K-ATPase channels. RhEPO pretreatment significantly inhibited serum cystatin-C (2 hr: 453 ± 64 μg/L vs 337 ± 28 μg/L, p < 0.01), serum neutrophil gelatinase-associated lipocalin (72 hr: 1,175 ± 107 ng/L vs 1,737 ± 402 ng/L, p < 0.05), and urinary fractional excretion of sodium (%) increase (0.9 ± 0.1 vs 2.2 ± 0.8, p < 0.05) and alleviated the decrease of urinary osmolality (1,293 ± 101 mosmol/kg H2O vs 767 ± 91 mosmol/kg H2O, p < 0.05) induced by ischemia-reperfusion injury. Meanwhile, recombinant human erythropoietin greatly improved the ischemia-reperfusion-induced attenuation of transient receptor potential channel-6 expression (48 hr: 42% ± 2% vs 67% ± 2% and 72 hr: 55% ± 2% vs 66% ± 2%), as well as aquaporin-2 and Na,K-ATPase expression in collecting ducts. Transient receptor potential channel-6 functionally interacted with Na,K-ATPase but not aquaporin-2.. Recombinant human erythropoietin pretreatment at the dose of 5,000 IU/kg potently prevented ischemia-reperfusion-induced acute tubular injury, which might be partly attributed to the restoring the effect of transient receptor potential channel-6 expression and collecting duct function.

Topics: Acute Kidney Injury; Acute-Phase Proteins; Animals; Aquaporin 2; Cystatin C; Erythropoietin; Humans; Kidney Tubules, Collecting; Lipocalin-2; Lipocalins; Male; Proto-Oncogene Proteins; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Sodium-Potassium-Exchanging ATPase; TRPC Cation Channels

Hypoxia/Reoxygenation (H/R) cardiac injury is of great importance in understanding Myocardial Infarctions, which affect a major part of the working population causing debilitating side effects and often-premature mortality. H/R injury primarily consists of apoptotic and necrotic death of cardiomyocytes due to a compromise in the integrity of the mitochondrial membrane. Major factors associated in the deregulation of the membrane include fluctuating reactive oxygen species (ROS), deregulation of mitochondrial permeability transport pore (MPTP), uncontrolled calcium (Ca2+) fluxes, and abnormal caspase-3 activity. Erythropoietin (EPO) is strongly inferred to be cardioprotective and acts by inhibiting the above-mentioned processes. Surprisingly, the underlying mechanism of EPO's action and H/R injury is yet to be fully investigated and elucidated. This study examined whether EPO maintains Ca2+ homeostasis and the mitochondrial membrane potential (ΔΨm) in cardiomyocytes when subjected to H/R injury and further explored the underlying mechanisms involved. H9C2 cells were exposed to different concentrations of EPO post-H/R, and 20 U/ml EPO was found to significantly increase cell viability by inhibiting the intracellular production of ROS and caspase-3 activity. The protective effect of EPO was abolished when H/R-induced H9C2 cells were treated with Wortmannin, an inhibitor of Akt, suggesting the mechanism of action through the activation Akt, a major survival pathway.

Topics: Animals; Caspase 3; Cell Hypoxia; Cell Line; Erythropoietin; Membrane Potential, Mitochondrial; Protective Agents; Rats; Reactive Oxygen Species; Reperfusion Injury; Signal Transduction

The parameters of blood oxygen transport, the products of lipid peroxidation and antioxidative factors were determined in rats during the hepatic ischemia/reperfusion (HIR) under preliminary single injections of different doses ofrecombinant human erythropoietin (rhEPO). Hepatic ischemia was induced for 30 min by Pringle maneuver, reperfusion lasted 120 min. The experiments had shown that HIR led to the significant decreases in the hemoglobin oxygen affinity, activation of lipid peroxidation, depletion of the antioxidant system, increases blood transaminases (ALT and AST). rhEPO in dose 100 IU/kg aggravates decreasing hemoglobin oxygen affinity, improves some antioxidant parameters, but didn't correct lipid peroxidation or plasma transaminases during HIR. rhEPO infusion id dose 1000 IU/kg leads to oxyhemoglobin dissociative curve shift left-wards, improves prooxidant-antioxidant balance and plasma ALT, AST activities at the end of re-perfusion period.

Topics: Alanine Transaminase; alpha-Tocopherol; Animals; Aspartate Aminotransferases; Blood Gas Analysis; Catalase; Erythropoietin; Hemoglobins; Lipid Peroxidation; Liver; Male; Rats; Reperfusion Injury; Schiff Bases; Vitamin A

Extensive research during the last decade demonstrated that a single systemic administration of -erythropoietin (EPO) lead to significant attenuation of myocardial infarction (MI) induced in animals, mostly small rodents, either by a myocardial ischemia followed by reperfusion or by a permanent ligation of a coronary artery. Both methods are critically reviewed with the aim of helping the reader in appreciating key issues in the translation of experimental results to the clinic. Results of several clinical trials in patients with acute MI completed to date failed to demonstrate beneficial effects of EPO, and thus put into question the validity of results obtained in animal models. Comprehensive review of design and results of animal experiments and clinical trials presented here allowed authors to postulate that therapeutic window for EPO during developing MI is very narrow and was possibly missed in negative clinical trials. This point was illustrated by the negative outcome of experiment in the rat model of MI in which timing of EPO administration was similar to that in clinical trials. The design of future clinical trials should allow for a narrow therapeutic window of EPO. Given current standards for onset-to-door and door-to-balloon time the optimal time for EPO administration should be just prior to PCI.

Topics: Animals; Erythropoietin; Humans; Myocardial Infarction; Myocytes, Cardiac; Reperfusion Injury

We investigated the effect of erythropoietin (EPO) on differentiation and secretion of bone marrow-derived mesenchymal stem cells in an acute kidney injury microenvironment. Acute kidney injury mouse models were prepared. Both renal cortices were then immediately collected to produce the ischemia/reperfusion kidney homogenate supernatant. The morphological and ultrastructural changes in the cells were observed using an inverted microscope and a transmission electron microscope. Cytokeratin-18 was detected using flow cytometry. Bone morphogenetic protein-7 levels, hepatocyte growth factor, and vascular endothelial growth factor in the culture medium were detected using an enzyme-linked immunosorbent assay. The cells had high CD29 and CD44 expression, as well as low CD34 and CD45 expression. More round and oval cells with cobble-like appearances were observed after EPO treatment. In addition, an increase in the number of rough endoplasmic reticula, lysosomes, and mitochondria was observed in the cytoplasm; the intercellular junction peculiar to epithelial cells was also seen on the cell surface. After treatment with ischemia/reperfusion kidney homogenate supernatant, cytokeratin-18 expression increased significantly and EPO could magnify its expression. Bone morphogenetic protein-7 levels, hepatocyte growth factor, and vascular endothelial growth factor levels after treatment with ischemia/reperfusion kidney homogenate supernatant significantly decreased, whereas EPO increased the cytokine secretion. The acute kidney injury microenvironment can induce the bone marrow-derived mesenchymal stem cells to partially differentiate into renal tubular epithelium-shaped cells, but weaken their secretion function. EPO intervention can boost up their differentiation function and reverse their low secretion effect.

Topics: Acute Kidney Injury; Animals; Antigens, CD34; Bone Marrow Cells; Bone Morphogenetic Protein 7; Cell Differentiation; Cells, Cultured; Erythropoietin; Hepatocyte Growth Factor; Hyaluronan Receptors; Integrin beta1; Keratin-18; Leukocyte Common Antigens; Male; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Inbred C57BL; Reperfusion Injury; Vascular Endothelial Growth Factor A

Erythropoietin (EPO) has been well recognized as a tissue protective agent by inhibiting apoptosis and inflammation. The tissue protective effect of EPO, however, only occurs at a high dosage, which may elicit severe side-effects at the meantime. Helix B surface peptide (HBSP), a novel peptide derived from the non-erythropoietic helix B of EPO, plays a specific role in tissue protection. We investigated effects of HBSP and the expression of its heterodimeric receptor, beta common receptor (βcR)/EPO receptor ( ), in a murine renal ischemia reperfusion (IR) injury model. HBSP significantly ameliorated renal dysfunction and tissue damage, decreased apoptotic cells in the kidney and reduced activation of caspase-9 and -3. The βcR/EPOR in the kidney was up-regulated by IR, but down-regulated by HBSP. Further investigation revealed that the expression and phosphorylation of Akt was dramatically enhanced by HBSP, but strongly reversed by wortmannin, the PI3K inhibitor. Wortmannin intervention improved βcR/EPOR expression, promoted caspase-9 and -3 activation, and increased active caspase-3 positive cells, while renal function and structure, and apoptotic cell counts scarcely changed. This result indicates a significant contribution of PI3K/Akt signaling pathway in the renoprotection of HBSP. The therapeutic effects of HBSP in this study suggest that HBSP could be a better candidate for renal protection.

Topics: Animals; Apoptosis; Erythropoietin; Ischemia; Kidney; Male; Mice; Mice, Inbred BALB C; Oncogene Protein v-akt; Peptides; Phosphatidylinositol 3-Kinases; Receptors, Erythropoietin; Reperfusion Injury; Signal Transduction

The exact mechanism by which erythropoietin protects the liver from ischemia reperfusion (I/R) injury is not yet known. In the present study, we examined the role of protein kinase B (PKB/AKT) and endothelial nitric oxide synthase (eNOS) in the protective effect of recombinant human erythropoietin (rHuEPO) on I/R injury of the liver.. We used a liver in situ I/R model. One hundred twenty adult male Sprague-Dawley rats were divided randomly into six groups. rHuEPO and (or) LY294002 were injected in the tail vein before the operation, and its effect was assessed by measuring the serum levels of aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, nitric oxide (NO), and endothelin-1 (ET-1) and by histologic analysis. The expression of erythropoietin receptor (EPOR) and eNOS was measured by real-time polymerase chain reaction. Total AKT and eNOS and phosphorylated AKT and eNOS were examined by western blot.. rHuEPO dramatically attenuated the functional and morphologic injuries. The serum levels of alanine aminotransferase and lactate dehydrogenase were significantly decreased, but the amount of NO in the serum was increased in the I/R + rHuEPO group. Accordingly, rHuEPO administration significantly ameliorated the histologic damages at 6 h after reperfusion. rHuEPO significantly stimulated the phosphorylation of AKT and eNOS in the rats after liver I/R.. The protective effect of rHuEPO in I/R injury is mediated via the activation of the phosphatidylinositol-3 kinase/AKT/eNOS signaling pathway, at least in part, by increasing p-AKT and p-eNOS and leads to the maintenance of an elevated level of NO.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Chromones; Disease Models, Animal; Enzyme Inhibitors; Erythropoietin; Humans; Ischemic Preconditioning; L-Lactate Dehydrogenase; Liver; Male; Morpholines; Nitric Oxide Synthase Type III; Phosphatidylinositol 3-Kinase; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Signal Transduction

To investigate the effect of recombine human erythropoietin (rhEPO) on apoptosis of neural cells in fetal rats after intrauterine hypoxic-ischemic injury.. Twenty SD rats on 19 days of pregnancy were divided into rhEPO (2500 U/kg, 5000 U/kg, 7500 U/kg) treated groups, ischemia-reperfusion (I/R) group and sham-operated group (4 rats in each group). Intrauterine hypoxic-ischemic injury of fetal rat was induced by bilateral occlusion of utero-ovarian artery for 20 min. rhEPO was injected into the rats in rhEPO treated group through the caudal vein 30 min before hypoxic-ischemic injury while saline was used in the other two groups. There was no hypoxic-ischemic injury in sham-operated group. The death rate of fetal rats was evaluated at 24 h after the operation, and then the brain samples of fetal rats were harvested. The expression of Caspase-3 protein was observed by immunohistochemistry. Neuroapoptosis was measured by TdT mediated dUTP-biotin nick end labeling (TUNEL) staining.. Death rates of fetal rats in rhEPO treated groups decreased compared with the I/R group (P < 0.05). Compared with the I/R group, there was less expression of copious Caspase-3 in rhEPO treated group (P < 0.01). The expression of Caspase-3 was decreased in the rhEPO treated groups with the increase of rhEPO dose (P < 0.01). Compared with the I/R group, the death rate of fetal rats in rhEPO treated groups decreased (P < 0.05), the number of apoptosis cells also decreased obviously (P < 0.01). The anti-apoptosis effect of 5000 U/kg rhEPO was similar to 7500 U/kg rhEPO, but better than 2500 U/kg rhEPO (P < 0.01).. rhEPO can inhibit the apoptosis of fetal rat brain cells after intrauterine hypoxic-ischemic injury.

Topics: Animals; Apoptosis; Brain; Caspase 3; Epoetin Alfa; Erythropoietin; Female; Fetus; Humans; Hypoxia; Neurons; Pregnancy; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury

SMND-309 is a novel derivative of salvianolic acid B, and has shown protective effects against rat cortical neuron damage in vitro and in vivo. However the molecular mechanisms through which SMND-309 affords this protection are unclear. The present study aimed to investigate the mechanisms associated with the protective activities of SMND-309 in a cerebral ischemia and reperfusion injury rat model. In this study, we used AG490, a specific inhibitor of the signaling pathway involving the Janus Kinase 2 (JAK2)/Signal Transducers and Activators of Transcription 3 (STAT3) signaling molecules and suramin, a potent inhibitor of vascular endothelial growth factor (VEGF), to investigate the mechanisms of SMND-309. The cerebral ischemia and reperfusion injury model was induced by performing middle cerebral artery occlusion (MCAO) in the rats. SMND-309 mitigated the effects of ischemia and reperfusion injury on brain by decreasing the infract volume, improving neurological function, increasing the survival of neurons and promoting angiogenesis by increasing the levels of erythropoietin (EPO), erythropoietin receptor (EPOR), phosphorylated JAK2 (P-JAK2), phosphorylated STAT3 (P-STAT3), VEGF and VEGF receptor 2 (Flk-1) in the brain. Our results suggest that SMND-309 provides significant neuroprotective effects against cerebral ischemia and reperfusion injury. The mechanisms of this protection may be attributed to the increased VEGF expression occurring from the JAK2/STAT3 pathway, activated by the increased EPO/EPOR expression in the brain.

Topics: Animals; Axons; Benzofurans; Brain; Brain Ischemia; Caffeic Acids; Cerebral Infarction; Dendrites; Erythropoietin; Gene Expression Regulation; Infarction, Middle Cerebral Artery; Janus Kinase 2; Male; Neovascularization, Physiologic; Neuroprotective Agents; Phosphoproteins; Platelet Endothelial Cell Adhesion Molecule-1; Rats; Rats, Sprague-Dawley; Receptors, Erythropoietin; Recovery of Function; Reperfusion Injury; Signal Transduction; STAT3 Transcription Factor; Survival Analysis; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2; Water

Cerebral ischemia-reperfusion injury can activate signal transducers and activators of transcription (STAT). STAT1 initiates neuronal apoptosis following cerebral ischemia-reperfusion, while STAT3 is neuroprotective. Erythropoietin (EPO) promotes regeneration through STAT3 and facilitates neuronal survival following ischemia. However, there are few reports on the effects of EPO on phosphorylated STAT1 (P-STAT1) level following cerebral ischemia-reperfusion in rats, and there is no evidence on the simultaneous observation of the four kinds of protein:STAT1, P-STAT1, STAT3, and P-STAT3.. We established a rat focal cerebral ischemia-reperfusion injury model, and used Western blot and immunohistochemical staining to assess the levels of STAT1 and STAT3 expression, and TdT-mediated dUTP-biotin nick end-labeling (TUNEL) was carried out to observe the number of apoptotic cells with or without EPO treatment.. Our findings show that EPO treatment had no significant effect on STAT1 and STAT3 expression, but P-STAT1 and P-STAT3 were slightly decreased and significantly increased, respectively, after EPO treatment. Neurologic deficits, the infarct volume, and the number of apoptotic cells were significantly decreased after EPO treatment.. The results suggest that EPO exerts a neuroprotective effect by influencing STAT3 and STAT1 expression in the area injured by cerebral ischemia-reperfusion.

Topics: Animals; Apoptosis; Blotting, Western; Epoetin Alfa; Erythropoietin; Hematinics; Immunohistochemistry; In Situ Nick-End Labeling; Infarction, Middle Cerebral Artery; Male; Neurologic Examination; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; STAT1 Transcription Factor; STAT3 Transcription Factor

To investigate the effects of recombine human erythropoietin (rhEPO) on neural cells apoptosis and the expression of Caspase-3 protein in brain tissue of fetal rats after intrauterine hypoxic-ischemic brain injury.. Forty-four Sprague-Dawley rats on 19 days of pregnancy were divided into rhEPO treated group, ischemia-reperfusion group and sham-operated group. Intrauterine hypoxic-ischemic injury of fetal rats was induced by bilateral occlusion of the utero-ovarian artery for 20 min. rhEPO (5000 U/kg) was injected into rats through caudal vein in rhEPO treated group while saline was injected into rats in hypoxic-ischemic group 30 min before hypoxic-ischemic injury. The brain samples in rhEPO treated group and hypoxic-ischemic group were obtained at 30 min, 3 h, 6 h, 24 h and 48 h respectively after artery clamping. There was no hypoxic-ischemic injury in sham-operated group, so the brain samples were obtained at 24 hours after sham operation. Neuroapoptosis in brain tissue was measured by TdT mediated dUTP-biotin nick end labeling (Tunel) staining. The expression of Caspase-3 protein was observed by immunohistochemistry.. The number of apoptosis cells in fetal rat hippocampus after intrauterine hypoxic-ischemic increased progressively with reperfusion. Compared with the I/R group, the number of apoptosis cells decreased in rhEPO treated group (P < 0.01). The expression of Caspase-3 increased rapidly after 3 hours from the reperfusion in the I/R group. Compared with the I/R group, there was less expression of Caspase-3 in rhEPO treated group (P < 0.01).. rhEPO showed the effects to inhibit the apoptosis of fetal neural cells and the expression of Caspase-3 protein due to intrauterine hypoxic-ischemic brain injury.

Topics: Animals; Brain; Caspase 3; Erythropoietin; Female; Fetal Hypoxia; Humans; Hypoxia-Ischemia, Brain; Ischemic Preconditioning; Male; Pregnancy; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury

Ischemia reperfusion (IR) and cyclosporine A (CsA) injuries are unavoidable in kidney transplantation and are associated with allograft dysfunction. Herein, the effect and mechanism of a novel tissue protective peptide, helix B surface peptide (HBSP) derived from erythropoietin, were investigated in a rat model. The right kidney was subjected to 45 min ischemia, followed by left nephrectomy and 2-week reperfusion, with or without daily treatment of CsA 25 mg/kg and/or HBSP 8 nmol/kg. Blood urea nitrogen was increased by CsA but decreased by HBSP at 1 week and 2 weeks, while the same changes were revealed in urinary protein/creatinine only at 2 weeks. HBSP also significantly ameliorated tubulointerstitial damage and interstitial fibrosis, which were gradually increased by IR and CsA. In addition, apoptotic cells, infiltrated inflammatory cells, and active caspase-3+ cells were greatly reduced by HBSP in the both IR and IR + CsA groups. The 17 kD active caspase-3 protein was decreased by HBSP in the IR and IR + CsA kidneys, with decreased mRNA only in the IR + CsA kidneys. Taken together, it has been demonstrated, for the first time, that HBSP effectively improved renal function and tissue damage caused by IR and/or CsA, which might be through reducing caspase-3 activation and synthesis, apoptosis, and inflammation.

Topics: Acute Kidney Injury; Animals; Apoptosis; Caspase 3; Cyclosporine; Enzyme Activation; Erythropoietin; Fibrosis; Gene Expression; Inflammation; Kidney; Kidney Function Tests; Male; Peptide Fragments; Protective Agents; Rats; Reperfusion Injury; RNA, Messenger

The effect of erythropoietin (EPO) on neonatal hearts is not well understood. The current hypothesis is that EPO has protective effects against ischaemia-reperfusion when administered prior to ischaemia induction.. Systolic and diastolic indices, as well as the Akt and extracellular-regulated kinase (Erk) signalling pathways, were studied in vivo using a neonatal pig heart model. Regional ischaemia was induced for 45 min by the ligation of the left anterior descending artery, followed by 90 min of reperfusion. The treatment groups consisted of: (i) untreated controls, (ii) treatment with EPO 3 min prior to ischaemia and (iii) treatment with EPO 24 h before ischaemia. Sophisticated myocardial contractility indices were assessed by pressure/volume loops of the left ventricle. The Akt and Erk pathways were evaluated via a western blot.. Elastance was found to be higher in the group receiving EPO 3 min prior to ischaemia. In addition, preload recruitable stroke work was higher for both groups receiving EPO prior to ischaemia when compared with controls. The time constant of the isovolumic relaxation and end-diastolic pressure-volume relationship did not differ between the three groups after 90 min of reperfusion. Furthermore, EPO treatment enhanced phosphorylation of Akt, but not Erk, and EPO-treated animals showed lower levels of apoptosis-related proteins.. EPO had a protective effect on neonatal systolic function after ischaemia/reperfusion injury, but no effect on diastolic function. This cardioprotective effect might be mediated by the activation of the Akt pathway.

Topics: Analysis of Variance; Animals; Animals, Newborn; Apoptosis; Cardiotonic Agents; Erythropoietin; Intracellular Signaling Peptides and Proteins; Male; Myocardial Contraction; Phosphorylation; Reperfusion Injury; Signal Transduction; Swine; Systole

Recombinant human erythropoietin (rHuEPO) is currently the mainstay of renal anaemia treatment. Recently, rHuEPO has been shown to provide pleiotrophic tissue protection in various pathological conditions. However, the benefits of rHuEPO beyond anaemia treatment are limited because it increases red blood cell mass. Carbamylated erythropoietin (CEPO) is the first rHuEPO derivative that lacks erythropoietic activity but retains tissue protection properties. Since carbamylation targets lysine residues on rHuEPo, we hypothesized that targeted lysine modifications of rHuEPO may result in a novel non-erythropoietic erythropoietin.. rHuEPO was subjected to various targeted lysine modifications. In vitro cytoprotection and apoptosis were evaluated using P19 and HEK293 cells. In vivo erythropoiesis was performed by administering the derivatives to animals for 2 weeks. Renoprotection was tested on an ischaemia/reperfusion (I/R) model.. We synthesized a novel derivative, a glutaraldehyde erythropoietin (GEPO). This construct abolished in vivo erythropoiesis. Biochemical characterization showed that GEPO was more electrostatically negative than rHuEPO. Immunoprecipitation experiments revealed that GEPO bound to the IL3RB/EPOR heterotrimeric receptor and ameliorated cellular apoptosis via the activation of Bcl-2. Notably, Bcl-2 activation was suppressed by the JAK2 inhibitor, tyrphostin AG490. In vivo experiments showed that GEPO also ameliorated kidney damage due to I/R injury both functionally and histologically.. Herein, we describe a novel lysine-modified rHuEPO, glutaradehyde-EPO (GEPO), obtained from a simple reaction. This derivative has no erythropoietic properties but retains cell-protective characteristics both in vitro and in vivo, with promise for future use as an adjunctive treatment of kidney disease.

Topics: Animals; Apoptosis; Cytoprotection; Erythrocyte Count; Erythropoiesis; Erythropoietin; Female; HEK293 Cells; Humans; Janus Kinase 2; Kidney; Mice; Protective Agents; Receptors, Erythropoietin; Recombinant Proteins; Reperfusion Injury

Administration of recombinant human erythropoietin (rhEPO) protects neurons from injury after brain ischemia-reperfusion (I/R), which is in part mediated by ameliorating the blood-brain barrier (BBB) leakage. But the mechanism of rhEPO's protective effects on BBB remains unclear. This study aims to investigate the effects of rhEPO on BBB integrity and the expressions of tight junctions (TJs) associated proteins of zonula occluden-1 (ZO-1), occludin, and claudin-5 in cerebral I/R rats. These rats underwent 2 h of ischemia and then were reperfused for up to 3 and 72 h. Animals were randomly divided into five groups: sham-operated group, I/R 3 h and I/R 72 h group (2 ml saline was injected intraperitoneally just before the onset of ischemia), rhEPO +I/R 3 h, and rhEPO +I/R 72 h group (5,000 U/kg rhEPO diluted in 2 ml saline solution was injected intraperitoneally just before the onset of ischemia). We verified that rhEPO could decrease the BBB leakage induced by I/R injury detected by Evans blue extravasation. 2, 3, 5-Triphenyltetrazolium chloride staining results showed that rhEPO decreased infarct volume after cerebral I/R. TJ integrity was partly restored by rhEPO observed by transmission electron microscopy. The mRNA and protein expression levels of ZO-1, occludin, and claudin-5 were significantly increased compared with I/R groups at the same reperfusion time point by reverse transcriptase-polymerase chain reaction and Western blot assays. The treatment of rhEPO induced the redistribution of ZO-1, occludin, and claudin-5 in cerebral microvessels observed by immunohistochemical staining. Compared with I/R groups, the mRNA level of tumor necrosis factor-alpha (TNF-α) in cerebral microvessels decreased markedly after rhEPO treatment, accompanied with reduced TNF-α protein level and nuclear factor-кB (NF-кB) p65 activation detected by enzyme-linked immunosorbent assay. These results suggested that the protective mechanism of rhEPO on BBB after cerebral I/R injury was associated with the upregulation of TJ-associated proteins. The downregulated TNF-α levels and NF-кB activation induced by rhEPO might be involved in this process.

Topics: Animals; Blood-Brain Barrier; Claudin-5; Erythropoietin; Male; Occludin; Rats; Rats, Sprague-Dawley; Reperfusion Injury; RNA, Messenger; Tight Junction Proteins; Tight Junctions; Transcription Factor RelA; Transcription, Genetic; Tumor Necrosis Factor-alpha; Zonula Occludens-1 Protein

To study the effect of erythropoietin (EPO) treatment on renal and lung injury following renal ischemia/reperfusion (I/R).. Thirty male Wistar rats were assigned to three groups of 10 rats each. The first group was sham-operated, the second was subjected to renal I/R (30 min of ischemia followed by 24 h of reperfusion). The third group was subjected to renal I/R and treated with EPO in two doses: the first dose 1 h prior to ischemia (1,000 U/kg) and the second dose 6 h after ischemia (1,000 U/kg).. The renal and lung tissue injury index, tissue serum blood urea nitrogen and creatinine (Cr) were higher in the renal I/R group compared to the renal I/R + EPO group; the difference was statistically significant (p < 0.05). Kidney and lung tissue glutathione peroxidase and superoxide dismutase levels were higher in the renal I/R + EPO group than the renal I/R group; the difference was also statistically significant (p < 0.05).. The data showed that EPO pretreatment could be effective in reducing renal and lung injury following renal I/R and could improve the cellular antioxidant defense system. Hence EPO pretreatment may be effective for attenuating renal and lung injury after renal I/R-induced injury during surgical procedures, hypotension, renal transplantation and other conditions inducing renal I/R.

Topics: Animals; Blood Urea Nitrogen; Creatinine; Disease Models, Animal; Erythropoietin; Glutathione Peroxidase; Kidney Diseases; Lung Diseases; Male; Oxidative Stress; Rats; Rats, Wistar; Reperfusion Injury; Superoxide Dismutase

Although ischemic preconditioning (IP) can provide powerful protection on brain against ischemic insult, it is rarely used in clinic to prevent the occurrence of ischemic stroke because of safety concerns. It is therefore necessary to seek the safer stimuli to initiate pharmacological preconditioning. Our previous work demonstrated that ginkgolide B (GB) could protect neurons against ischemia-induced apoptosis. Astrocytes are the most numerous cells in mammalian central nervous system and there is a close bi-directional communication between neurons and astrocytes in brain. Besides neurons, whether GB can exert the role of preconditioning on astrocytes through which to further improve neuronal survival under ischemic condition is not yet known. In the present study, primary cultured astrocytes were treated with GB for 24h or short-term ischemia (ischemia for 3h, as ischemic preconditioning/IP), and then cultured back to normoxia and normal medium for 24h to induce the preconditioning response. Astrocyte-conditioned medium (ACM) was then collected and used to incubate the cultured neurons for 24h before neurons were subjected to severe ischemia. Our results demonstrated that not only GB and IP increased astrocytic viability in ischemia, but also the conditioned medium from astrocytes treated with GB or IP increased cell viability and decreased the number of apoptosis of neurons in ischemia. We also found that GB and IP significantly stimulated astrocytes to express and secrete erythropoietin (EPO) into ACM, and the addition of anti-EPO antibody blocked the protective effect of GB or IP-treated astrocytes culture medium on neurons in ischemia. Further study of above protection revealed that ACM from astrocytes treated with GB or IP induced the inactivation of proapoptotic factor Bad by phosphorylation at serine 136 and 112 ((136)p-Bad and (112)p-Bad) in neurons. Together, our results suggest that GB is capable of preconditioning on astrocytes as IP and then protects neurons against ischemia-induced apoptosis, which is mediated by EPO.

Topics: Animals; Astrocytes; Blotting, Western; Enzyme-Linked Immunosorbent Assay; Erythropoietin; Ginkgolides; Ischemic Preconditioning; Lactones; Mice; Mice, Inbred ICR; Neurons; Reperfusion Injury; Up-Regulation

To test the hypothesis that a carbamylated EPO-FC fusion protein (cEPO-FC) or recombinant human erythropoietin (rhEPO) would protect against kidney ischemia/reperfusion (I/R) injury in pigs with atherosclerosis.. Anesthetized and mechanically ventilated animals received cEPO-FC (50 μg kg(-1)), rhEPO (5,000 IU kg(-1)), or vehicle (n = 9 per group) prior to 120 min of aortic occlusion and over 4 h of reperfusion. During aortic occlusion, mean arterial pressure (MAP) was maintained at 80-120 % of baseline values by esmolol, nitroglycerin, and ATP. During reperfusion, noradrenaline was titrated to keep MAP at pre-ischemic levels. Blood creatinine and neutrophil gelatinase-associated lipocalin (NGAL) levels, creatinine clearance, fractional Na(+) excretion, and HE and PAS staining were used to assess kidney function and histological damage. Plasma interleukin-6, tumor necrosis factor-α, nitrate + nitrite and 8-isoprostane levels were measured to assess systemic inflammation, and nitrosative and oxidative stress.. I/R caused acute kidney injury with reduced creatinine clearance, increased fractional Na(+) excretion and NGAL levels, moderate to severe glomerular and tubular damage and apoptosis, systemic inflammation and oxidative and nitrosative stress, but there were no differences between the treatment groups. Pre-ischemia nitrate + nitrite and 8-isoprostanes levels were lower and higher, respectively, than in healthy animals of a previous study, and immune histochemistry showed higher endothelial nitric oxide synthase and lower EPO receptor expression in pre-ischemia kidney biopsies than in biopsies from healthy animals.. In swine with atherosclerosis, rhEPO and cEPO-FC failed to attenuate prolonged ischemia-induced kidney injury within an 8-h reperfusion period, possibly due to reduced EPO receptor expression resulting from pre-existing oxidative stress and/or reduced NO release.

Topics: Animals; Erythropoietin; Immunoglobulin G; Kidney; Male; Recombinant Proteins; Reperfusion Injury; Swine

Organ shortage in liver transplantation has justified usage of marginal donor livers to expand the donor organ pool. The particular susceptibility of steatotic livers to I/R injury necessitates optimal preservation conditions in order to minimize preservation-reperfusion injury for successful transplantation.. The effect of erythropoietin (EPO) as additive to HTK preservation solution was studied in a mouse model. Lean and steatotic livers were harvested, stored for 24 h in 4°C HTK solution containing either EPO or saline and reperfused for 2 h with 37°C Krebs-Henseleit buffer. Livers without cold storage served as sham controls.. Flushing of livers upon cold storage revealed a transaminase release, which was 2- to 10-fold higher in steatotic versus lean livers. EPO was effective in reducing the enzyme release to 50% in steatotic but not in lean livers. EPO prevented cold storage-induced denudation of the endothelial lining in steatotic livers, but aggravated it in lean livers. During reperfusion, steatotic livers presented with lower oxygen consumption and higher enzyme release than lean livers. In all livers, parameters of reperfusion injury remained unaffected by EPO. Expression of UCP2 was found markedly higher in steatotic livers. After I/R, steatotic livers revealed a significant drop of UCP2, whereas expression in lean livers was only slightly affected. EPO diminished Erk phosphorylation to almost the same extent in both mouse strains.. Fortification of the preservation solution by EPO ameliorates cold ischemic injury of steatotic livers and may thus be considered for use as an adjunctive agent to increase the success of transplanting steatotic livers.

Topics: Animals; Cold Temperature; Disease Models, Animal; Erythropoietin; Fatty Liver; Female; Glucose; Ion Channels; Liver; Liver Transplantation; Male; Mannitol; Mice; Mice, Inbred Strains; Mice, Obese; Mitochondrial Proteins; Mitogen-Activated Protein Kinase Kinases; Obesity; Organ Preservation Solutions; Potassium Chloride; Procaine; Receptors, Erythropoietin; Reperfusion Injury; Signal Transduction; Thinness; Uncoupling Protein 2

Inhibition of the HIF regulating prolyl hydroxylation domain (PHDs) proteins prior to renal injury (preconditioning) has been shown to protect the kidney via activation of hypoxia-inducible transcription factors (HIF). Application of erythropoietin (EPO), one of the HIF target genes, has also been shown to be nephroprotective, and it remains unclear to what extent the effect of HIF induction is mediated by EPO. It is also unknown whether HIF activation after the onset of ischaemia (postconditioning) is still able to protect the kidney.. Using a rat model of renal ischaemia-reperfusion injury, animals were treated with the PHD inhibitor (PHD-I) 2-(1-chloro-4-hydroxyisoquinoline-3-carboxamido) acetate (ICA), vehicle (Veh) or recombinant human EPO (300 IU/kg) 6 h (ICA or Veh) or 30 min (EPO) prior to ischaemia (preconditioning) or with ICA prior to reperfusion (postconditioning). Renal function was assessed at baseline, 24 h and 72 h. After 72 h, kidneys were processed for histology and morphometric analysis. HIF immunohistochemistry and real-time polymerase chain reaction for HIF target genes, including EPO, were performed to evaluate ICA effects.. ICA treatment resulted in stabilization of HIF-1α and -2α and up-regulation of HIF target genes in a dose-dependent manner. Preconditional activation of HIF by ICA significantly improved serum creatinine levels and renal morphology in comparison to Veh (P < 0.05), while postconditional ICA treatment was ineffective. EPO therapy improved tissue morphology but had no impact on the course of serum creatinine.. These findings are in line with the concept that PHD-Is exert their protective effects through accumulation of HIF target gene products, with time requirements for increased transcription and translation of HIF-dependent genes, and suggest that their renoprotective effect is not predominately mediated by EPO.

Topics: Acute Kidney Injury; Animals; Enzyme Inhibitors; Epoetin Alfa; Erythropoietin; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Immunoenzyme Techniques; Male; Procollagen-Proline Dioxygenase; Rats; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; Recombinant Proteins; Reperfusion Injury; RNA, Messenger

Tubulointerstitial inflammation is the characteristics of renal ischemia reperfusion injury (IRI) that is inevitable in kidney transplantation. Erythropoietin (EPO) has recently been shown to have protective effects on renal IRI by anti-apoptosis and anti-oxidation. Here, the effect and mechanism of EPO on renal IRI were further investigated, with a focus on tubulointerstitial inflammation.. Male Sprague-Dawley rats were administrated with saline or EPO prior to IRI induced by bilateral renal pedicle clamping. Twenty-four hours following reperfusion, the effects of EPO on renal IRI were assessed by renal function and structure, tubulointerstitial myeloperoxidase (MPO) positive neutrophils, and proinflammatory mediator gene expression. The translocation and activity of NF-κB in renal tissues were also evaluated.. Compared with control groups, the EPO treated group exhibited lower serum urea and creatinine levels, limited tubular necrosis with a lower score of renal histological lesion. MPO positive cells in the tubulointerstitial area were greatly increased by IRI, but significantly reduced by the treatment of EPO. The gene expression of proinflammatory cytokines (IL-1β, IL-6, IL-10, and TNF-α) and chemokine (MCP-1) was also significantly decreased by EPO. In addition, less activation and nuclear-translocation of NF-κB was observed in the kidney treated by EPO as well.. EPO improved renal function and structure in IRI rats via reducing neutrophils in the tubulointerstitium, the production of proinflammatory cytokines and chemokine, as well as the activation and nuclear-translocation of NF-κB. EPO may have potential clinical applications as an anti-inflammation agent clinically for a wide range of injury.

Topics: Animals; Apoptosis; Creatinine; Cytokines; Erythropoietin; Kidney; Male; Models, Animal; Necrosis; Nephritis, Interstitial; NF-kappa B; Oxidative Stress; Peroxidase; Rats; Rats, Sprague-Dawley; Reperfusion Injury

In pre-clinical studies, acute erythropoietin (EPO) administration has been shown to mitigate the deleterious effects of ischemia/reperfusion injury. We reviewed our clinical experience with intraoperative EPO administration as a potential renoprotective agent during laparoscopic partial nephrectomy (LPN).. Patients who underwent LPN at our institution between August 2008 and March 2010 received 500 IU/kg EPO 30 min prior to hilar occlusion. Those who underwent LPN between August 2006 and July 2008 without receiving EPO were selected as controls. Demographic, clinical, perioperative, and estimated glomerular filtration rate (eGFR) data were compared for the cohorts preoperatively, and during short-term (<6 months) and long-term (≥6 months) follow-up.. Short-term eGFR was evaluable for 39 EPO and 29 controls, while long-term eGFR was evaluable for 26 EPO and 27 controls. Baseline demographic and clinical features of the cohorts were similar. For EPO versus controls, median short and long-term follow-up was 19 days versus 22 days and 10.2 months versus 11.9 months, respectively. Mean preoperative, postoperative, and % change in eGFR were statistically similar for the cohorts during short- and long-term follow-up, without and with adjustment for baseline renal function (unadjusted P-values = 0.28, 0.095, and 0.38, respectively, short term, and 0.61, 0.50, and 0.69, respectively, long term).. In this retrospective study, a single dose of EPO prior to hilar occlusion during LPN had no added protective impact on postoperative eGFR in the short or long term. Prospective evaluation in patients with solitary kidneys may better elucidate its potential renoprotective role in this setting.

Topics: Adult; Aged; Cohort Studies; Creatinine; Dose-Response Relationship, Drug; Erythropoietin; Female; Follow-Up Studies; Glomerular Filtration Rate; Humans; Intraoperative Period; Kidney; Laparoscopy; Male; Middle Aged; Minimally Invasive Surgical Procedures; Nephrectomy; Postoperative Period; Renal Insufficiency, Chronic; Reperfusion Injury; Retrospective Studies; Treatment Outcome; Warm Ischemia

Ischemia-reperfusion (I/R) cannot be avoided in liver transplantation procedures, and apoptosis is a central mechanism of cell death after liver reperfusion. Protective effect of recombinant erythropoietin (rhEPO) on liver apoptosis has not been clearly investigated. This work investigated intraportal (IP) rhEPO-protective effect in a rat model of hepatic I/R-induced apoptosis and its appropriated time and dose of administration. Eight groups were included (n = 10/group): sham-operated, I/R (45 min ischemia and 2 h reperfusion), preconditioned rhEPO I/R (24 h or 30 min before ischemia), and postconditioned rhEPO I/R (before reperfusion) using two different rhEPO doses (1,000 and 5,000 IU/kg). When compared with the sham-operated group, the I/R group showed significant increase of serum levels of aspartate and alanine aminotransferases (AST, ALT), hepatic caspase-9 activity(894.99 ± 176.90 relative fluorescence units (RFU)/mg/min versus 458.48 ± 82.96 RFU/mg/min), and Fas ligand (FasL) expression, histopathological damages, and significant decrease in the antiapoptotic Bcl-xL/apoptotic Bax ratio(0.38 ± 0.21 versus 3.35 ± 0.77) rhEPO-improved ALT and AST but failed to reduce FasL expression in all groups compared with the I/R group. Thirty minutes and 24 h preconditioning with rhEPO (1,000 IU/kg) increased Bcl-xL/Bax ratio and reduced caspase-9 activity, and the same effect was observed when higher dose was given 24 h before ischemia. Preconditioning was more effective than postconditioning in improving caspase-9 activity, and no dose-dependent effect was observed. In conclusion, single IP rhEPO injection 30 min before ischemia has an advantage over rhEPO postconditioning in improving post-hepatic I/R-induced apoptosis with no additional time- and dose-dependent effects which may provide potentially useful guide in liver transplantation procedures.

Topics: Alanine Transaminase; Animals; Apoptosis; Apoptosis Regulatory Proteins; Aspartate Aminotransferases; Caspase 9; Enzyme Activation; Erythropoietin; Fas Ligand Protein; Gene Expression Regulation; Humans; Ischemia; Ischemic Postconditioning; Ischemic Preconditioning; Liver; Male; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury; Transcription, Genetic

Topics: Animals; Erythropoietin; Kidney; Male; Nephritis, Interstitial; Reperfusion Injury

Experimental study.. To investigate the effect of erythropoietin (EPO) pretreatment on spinal cord ischemic injury.. Experimental Research Center at Seoul National University Bundang Hospital, Korea.. Rats were treated with either 1000 IU kg(-1) of EPO (EPO group, n=8) or saline (control group, n=8) 24 h before ischemia. Spinal cord ischemia was induced using a balloon-tipped catheter placed on the proximal descending aorta in the control group and the EPO group, but not in the sham group (n=8). Neurological function was assessed using the motor deficit index (MDI; 0=normal, 6=complete paralysis) until 7 days after reperfusion, and histological examination of spinal cord was performed.. At the first day after reperfusion, the EPO group demonstrated a significantly lower MDI compared with the control group (2.0 (0.3-2.0) vs 4.0 (3.0-4.8), median (interquartile range); EPO group vs control group, respectively; P=0.002). This trend was sustained until 7 days after reperfusion (1.0 (1.0-1.8) vs 4.5 (3.3-5.0); EPO group vs control group, respectively; P=0.001), and more normal motor neurons (29.9±3.1 vs 21.4±3.4, mean±s.d.; EPO group vs control group, respectively; P<0.001) were observed. However, compared with the sham group, the EPO group displayed a significantly higher MDI (0.0, sham group) and fewer intact motor neurons (37.8±5.5, sham group; P<0.001, sham vs control group).. Pretreatment with EPO significantly attenuates neurological injury following spinal cord ischemia, although it cannot completely abolish the ischemic injury.

Topics: Animals; Disease Models, Animal; Erythropoietin; Male; Motor Neurons; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Spinal Cord Ischemia

It has been reported that erythropoietin protects the kidneys from ischemia/reperfusion injury. In the present study, we examined the role of Akt and endothelial nitric oxide synthase in the protective effect of erythropoietin on ischemia/reperfusion injury of the kidney.. Erythropoietin was injected in the peritoneal space of ICR mice after ischemia/reperfusion injury and its effect was assessed by measuring blood urea nitrogen and creatinine, and by histological analysis. Phosphorylation of Akt and endothelial nitric oxide synthase was examined by western blot analysis. Endothelial nitric oxide synthase gene null mice were also used to examine the role of endothelial nitric oxide synthase in the renoprotective effect of erythropoietin.. Erythropoietin administration significantly inhibited the increase in blood urea nitrogen and creatinine after ischemia/reperfusion injury compared with control mice. Accordingly, erythropoietin administration significantly ameliorated the histological damages, including apoptotic cell death. Erythropoietin significantly stimulated phosphorylation of Akt and endothelial nitric oxide synthase in the kidneys. When endothelial nitric oxide synthase gene null mice were subjected to ischemia/reperfusion injury, erythropoietin did not significantly suppress the increase in blood urea nitrogen or creatinine.. Erythropoietin seems to activate the Akt/endothelial nitric oxide synthase-dependent pathway in the kidneys. This pathway might be implicated in the renoprotective effect of erythropoietin in the ischemia/reperfusion injury model.

Topics: Animals; Apoptosis; Blood Urea Nitrogen; Creatinine; Erythropoietin; Kidney; Male; Mice; Mice, Inbred C57BL; Mice, Inbred ICR; Mice, Knockout; Nitric Oxide Synthase Type III; Phosphorylation; Proto-Oncogene Proteins c-akt; Reperfusion Injury; Signal Transduction

Topics: Animals; Erythropoietin; Kidney; Male; Nitric Oxide Synthase Type III; Proto-Oncogene Proteins c-akt; Reperfusion Injury

Pyruvate is known to be cytoprotective through antioxidant and anti-inflammatory mechanisms. We tested the hypothesis that pyruvate protects the brain against ischemia-reperfusion injury by inducing endogenous erythropoietin (EPO) expression.. Pyruvate's protective effect was evaluated in C6 glioma cells and HT22 neuronal cells subjected to transient oxygen glucose deprivation. Cell viability (calcein AM assay) and expression of hypoxia-inducible factor-1α, EPO, Akt and Erk (immunoblot), and EPO receptor (reverse transcription-polymerase chain reaction) were analyzed. Transient focal cerebral ischemia in rats was induced by 2 hours middle cerebral artery occlusion followed by 24 hours reperfusion. Pyruvate or saline was infused from 60 minutes occlusion until 30 minutes reperfusion. Lesion volume and DNA fragmentation were assessed by 2,3,5-triphenyltetrazolium staining and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling assay, respectively. Immunoblots were conducted to determine cerebral EPO contents.. Pyruvate increased cell viability, hypoxia-inducible factor-1α, EPO, and Akt phosphorylation. Small interfering RNA suppression of hypoxia-inducible factor-1α and EPO abolished pyruvate-induced cytoprotection. In the rat stroke model, pyruvate reduced lesion volume by 84% and DNA fragmentation by 77% versus controls; increased EPO content paralleled these cerebroprotective actions of pyruvate.. Pyruvate activation of the hypoxia-inducible factor-1α-EPO signaling cascade in neurons and glia could protect the brain from ischemia-reperfusion injury.

Topics: Animals; Brain; Cell Line, Tumor; Disease Models, Animal; Erythropoietin; Gene Expression Regulation; Hypoxia-Inducible Factor 1, alpha Subunit; Male; Mice; Proto-Oncogene Proteins c-akt; Pyruvic Acid; Rats; Rats, Sprague-Dawley; Receptors, Erythropoietin; Reperfusion Injury; Signal Transduction; Stroke

Cerebral ischemia-reperfusion (I/R) injury induces secondary cerebral damage. As drugs for treating this type of injury have shown poor efficacy and adverse side effects in clinical trials, a novel therapeutic strategy has been long awaited. In this study, we focused on the disruption of the blood-brain barrier after stroke, and applied a liposomal drug delivery system (DDS) designed to enhance the pharmacological effect of the neuroprotectant and to avoid side effects. PEGylated liposomes were injected at varying time after the start of reperfusion in transient middle cerebral artery occlusion (t-MCAO) model rats. The results showed PEGylated liposomes accumulated in the ischemic hemisphere at an early stage after reperfusion and were retained in the lesion for at least 24h after injection. We also investigated the effectiveness of asialo-erythropoietin (AEPO)-modified PEGylated liposomes (AEPO-liposomes) in treating the cerebral I/R injury. AEPO-liposome treatment significantly reduced TTC-defined cerebral legion following cerebral I/R injury, and ameliorated motor function compared with vehicle and AEPO treatment. In conclusion, these results indicate that AEPO-liposomes are a promising liposomal formulation for protecting the brain from I/R injury, and that this liposomal DDS has potential as a novel strategy for the treatment of cerebral I/R injury.

Topics: Animals; Apoptosis; Asialoglycoproteins; Blood-Brain Barrier; Cell Culture Techniques; Cell Survival; Disease Models, Animal; Drug Delivery Systems; Erythropoietin; In Situ Nick-End Labeling; Ischemic Attack, Transient; Liposomes; Male; Neuroprotective Agents; PC12 Cells; Rats; Rats, Wistar; Reperfusion Injury; Tissue Distribution

Darbepoetin-alpha (DA) is a novel erythropoiesis-stimulating agent developed for treating anemia. In animal models, recombinant human erythropoietin has been reported to be beneficial for neuroprotection. In this study, we determined whether DA would protect the spinal cord against ischemia-reperfusion injury in a rabbit model.. Forty rabbits were randomized into five groups of eight animals each: group 1 (sham), group 2 (ischemia), group 3 (vehicle), group 4 (30 mg/kg methylprednisolone), group 5 (30 μg/kg DA). Only laparotomy was performed in the sham group. In all the other groups, the spinal cord ischemia model was created by a 20-min occlusion of the aorta just caudal to renal artery with an aneurysm clip. The drugs were administered immediately after the clamp was removed. The animals were killed 24 h later. Spinal cord segments between L2 and L5 were harvested for analysis. Neurological evaluation was performed with the Tarlov scoring system just before the animals were killed. Level of tissue malondialdehyde was analyzed as a marker of lipid peroxidation and tissue caspase-3 activity as a marker of apoptosis. Also, histopathological evaluation of the tissues was performed.. Both malondialdehyde and caspase-3 levels were significantly decreased by DA administration. Histopathological evaluation of the tissues also demonstrated decrease in neuronal degeneration and infiltration parameters after DA administration. In the DA group, neurological outcome scores were statistically significantly better compared with the ischemia and the vehicle groups.. Although further studies considering different dose regimens and time intervals are required, DA was shown to be at least as effective as methylprednisolone in spinal cord ischemia/reperfusion model.

Topics: Anemia; Animals; Darbepoetin alfa; Disease Models, Animal; Erythropoietin; Hematinics; Male; Neuroprotective Agents; Rabbits; Reperfusion Injury; Spinal Cord Ischemia

The introduction of cyclosporine (CsA) has improved graft survival, but it causes nephropathy, which limits its clinical utility. Recently, we reported that carbamylated erythropoietin (CEPO) protected kidneys from ischemia reperfusion injury as well as EPO. To investigate the clinical applications of CEPO, we next evaluated the long-term therapeutic effect of CEPO using a CsA-induced nephropathy model. CsA caused renal dysfunction, while EPO/CEPO administration significantly improved renal function. EPO treatment significantly increased Hb concentration, while CEPO treatment neither enhanced nor reduced Hb concentration. CsA treatment induced tubular apoptosis, while EPO/CEPO administration inhibited it and increased PI3 kinase activation and Akt phosphorylation. In parallel, morphological assessment revealed that EPO/CEPO significantly reduced CsA-induced interstitial fibrosis and inhibited interstitial macrophage infiltration. In addition, real-time RT-PCR demonstrated that cortical mRNA levels of TGF-β1 and type I collagen were suppressed in the EPO/CEPO group. These results suggest a new therapeutic approach using CEPO to protect kidneys from CsA-induced nephropathy.

Topics: Animals; Apoptosis; Collagen Type I; Cyclosporine; Disease Models, Animal; Dogs; Erythropoiesis; Erythropoietin; Fibrosis; Hemoglobins; Humans; Immunosuppressive Agents; Kidney; Kidney Diseases; Macrophages; Male; Phosphatidylinositol 3-Kinases; Phosphorylation; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction; Transforming Growth Factor beta1

Interest has recently been focused on the possible role of bone marrow-originating stem cells and the therapeutic role of erythropoietin in the recovery of ischemia-induced acute kidney injury (AKI). The aim of the present study was to compare treatment with mesenchymal stem cells (MSCs) to treatment with darbepoetin-α (DPO) or both concomitantly in a rat model of ischemia/reperfusion (I/R) AKI.. Forty male Sprague-Dawley rats were included, and 28 of them were randomly assigned to controls (treated with serum physiologic) or one of the three treatment groups treated with either DPO, MSCs, or both (MSCs and DPO concomitantly) after the induction of I/R injury. Hematocrit, serum creatinine, and BUN levels were obtained at 0, 24, 48, and 72 h of surgery, and renal tissue was obtained at 72 h after nephrectomy for histological analysis. Tissue injury was quantified by standardized histological scoring systems, using light and electron microscopes.. Treatment with MSCs or DPO improved renal function compared with controls. However, the improvement observed in renal function in the MSC/DPO group was better than that in the other groups. Histological analysis demonstrated that tissue injury was significantly decreased in rats in the MSC or DPO groups compared to that of the controls; however the best recovery was observed in rats treated with MSCs and DPO concomitantly.. These results suggest that concomitant application of DPO and MSCs may be a potential novel renoprotective therapy for patients after having sustained an ischemic renal insult.

Topics: Acute Kidney Injury; Animals; Apoptosis; Blood Urea Nitrogen; Combined Modality Therapy; Creatinine; Darbepoetin alfa; Erythropoietin; Hematinics; Hematocrit; Ischemia; Kidney; Male; Mesenchymal Stem Cell Transplantation; Random Allocation; Rats; Rats, Sprague-Dawley; Reperfusion Injury

The tissue-protective effects of erythropoietin (EPO) have been extensively investigated, and EPO administration can raise the hemoglobin (Hb) concentration. Recently, we reported that carbamylated erythropoietin (CEPO) protected kidneys from ischemia-reperfusion injury as well as EPO.. To investigate the clinical applications of CEPO, we next evaluated the long-term therapeutic effect of CEPO using a tubulointerstitial model rat. We randomized remnant kidney model rats to receive saline, EPO, or CEPO for 8 weeks.. CEPO- and EPO-treated rats had improved serum creatinine levels compared with saline-treated remnant kidney model rats, although the Hb level was significantly increased in EPO-treated rats. Two-photon microscopy revealed that EPO/CEPO significantly ameliorated tubular epithelial cell damage assessed by endocytosis. In addition, CEPO or EPO protected endothelial cells with a sustained blood flow rate. EPO or CEPO suppressed the number of TUNEL-positive apoptotic cells with weak αSMA staining. Furthermore, PCR analysis demonstrated that TGF-β and type I collagen expression was attenuated in EPO- or CEPO-treated rats, accompanied by a significant decrease in interstitial fibrosis.. We established a long-term therapeutic approach to protect tubulointerstitial injury with CEPO, and thus, the therapeutic value of this approach warrants further attention and preclinical studies.

Topics: Animals; Collagen Type I; Creatinine; Disease Models, Animal; Erythropoietin; Fibrosis; Hemoglobins; Kidney; Kidney Diseases; Male; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Transforming Growth Factor beta

Acute renal failure (ARF) can be caused by injuries that induce tissue hypoxia, which in turn can trigger adaptive or inflammatory responses. We previously showed the participation of basic fibroblast growth factor (FGF-2) in renal repair. Based on this, the aim of this study was to analyze the effect of FGF-2 signaling pathway manipulation at hypoxia-induced protein levels, as well as in key proteins from the vasoactive systems of the kidney. We injected rat kidneys with FGF-2 recombinant protein (r-FGF) or FGF-2 receptor antisense oligonucleotide (FGFR2-ASO) after bilateral ischemia, and evaluated the presence of iNOS, EPO and HO-1, in representation of hypoxia-induced proteins, as well as COX-2, renin, kallikrein, and B2KR, in representation of the vasoactive systems of the kidney. A reduction in iNOS, HO-1, EPO, renin, kallikrein, B2KR, and in renal damage was observed in animals treated with r-FGF. The opposite effect was found with FGF-2 receptor down-regulation. In contrast, COX-2 protein levels were higher in kidneys treated with r-FGF and lower in those that received FGFR2-ASO, as compared to saline treated kidneys. These results suggest that the protective role of FGF-2 in the pathogenesis of ARF induced by I/R is a complex process, through which a differential regulation of metabolic pathways takes place.

Topics: Acute Kidney Injury; Animals; Cell Hypoxia; Cyclooxygenase 2; Disease Models, Animal; Erythropoietin; Fibroblast Growth Factor 2; Heme Oxygenase-1; Kallikreins; Kidney; Male; Nitric Oxide Synthase; Rats; Rats, Sprague-Dawley; Receptor, Bradykinin B2; Reperfusion Injury

To study whether recombinant human erythropoietin (rhEPO) reduces neuronal apoptosis through inhibiting over-expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in nucleus induced by brain ischemia/reperfusion in rats, 48 adult Sprague-Dawley rats were randomly divided into 3 groups: sham, saline and EPO groups. Animal models of brain ischemia/reperfusion were established by middle cerebral artery occlusion in rats. The effects of EPO on the sizes of ischemia tissue were observed by TTC staining. The over-expression of GAPDH in nucleus was detected by Hoechst-33258 and anti-GAPDH antibody double staining. The neuronal apoptosis in penumbral was detected by Nissl's staining and Hoechst-33258 immunofluorescence, respectively. The results showed that rhEPO treatment (3 000 U/kg, three times daily, i.p.) apparently reduced the sizes of infarct brain tissue in ischemia/reperfusion rats. rhEPO inhibited over-expression of GAPDH in nucleus of apoptotic neurons. In the meantime rhEPO decreased the number of apoptotic neurons in ischemia/reperfusion rats. These results suggest that rhEPO may induced reduction of neuronal apoptosis in penumbra may be through inhibiting over-expression of GAPDH in nucleus of apoptotic neurons induced by ischemia/reperfusion. Reduction of GAPDH over-expression in nucleus may play a pivotal role in EPO inhibiting neuronal apoptosis in cerebral ischemia/reperfusion rats, providing experimental evidence for EPO neuro-protecting effects against ischemia/reperfusion.

Topics: Animals; Apoptosis; Brain; Brain Ischemia; Erythropoietin; Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating); Humans; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury

Modification of the liposomal surface with a targeting molecule is a promising approach for the targeted delivery of therapeutics. Asialo-erythropoietin (AEPO) is a potent tool for targeting an ischemic region by binding to the EPO receptors on neuronal cells. Additionally, it shows a strong cytoprotective effect against programed cell death. Hence, AEPO-modified liposomes appear likely to have both a neuronal-targeting character and a neuroprotective effect on cerebral ischemic injury. In this study, we assessed the targeting ability of AEPO-modified PEGylated liposomes (AEPO-liposomes) to ischemic region and their improvement effect on neurological deficits induced by ischemia/reperfusion (I/R) in transient middle cerebral artery occlusion (t-MCAO) rats. Immunohistological analysis showed that the AEPO-liposomes given immediately after reperfusion extravasated into the ischemic region and attached strongly to neuronal cells. Also, neuronal nuclei (NeuN) staining was clearly visible only in the AEPO-liposome-treated group, suggesting that AEPO-liposomes protected neuronal cells from ischemia/reperfusion-induced damage. Moreover, a single administration of low-dose AEPO-liposomes significantly improved the neurological deficit compared to vehicle and free-AEPO treatment at 7 days after injection. In conclusion, AEPO-liposomes have clear potential as a neuroprotectant after stroke and as a DDS device targeting ischemic regions.

Topics: Animals; Asialoglycoproteins; Erythropoietin; Infarction, Middle Cerebral Artery; Liposomes; Male; Motor Activity; Neurons; Neuroprotective Agents; Polyethylene Glycols; Rats; Rats, Wistar; Reperfusion Injury

Carbamylated erythropoietin (C-EPO), one of the erythropoietin derivatives, retains strong anti-edema and neuroprotective properties while lacking the hematopoietic complications of erythropoietin. This study investigated the intracellular and molecular mechanisms underlying the anti-edema property of C-EPO. An in vitro model of astrocyte swelling was created by 5h of oxygen-glucose deprivation and subsequent reperfusion (OGD/Rep). Astrocyte cultures were then treated with C-EPO or left as control cells. Here we show that increases in astrocyte volume, morphological cell swelling, and changes in ultrastructure after OGD/Rep were significantly mitigated by treatment with C-EPO (10 ng/ml). The decreases in AQP-4 phosphorylation after OGD/Rep were remarkably recovered by C-EPO treatment. The OGD/Rep-induced upregulations of AQP-4 mRNA and protein were also prevented by C-EPO treatment. Additional treatment with phorbol myristate acetate, an activator of protein kinase C (PKC), enhanced C-EPO-mediated neuroprotective effects, while that of H-7, an inhibitor of PKC, blocked these protections. Our findings establish that C-EPO effectively mitigates astrocyte swelling induced by ischemia and reperfusion-like injury. The modulation of AQP-4 phosphorylation and expression via the PKC pathway is participated in the neuroprotective effects of C-EPO.

Topics: Animals; Animals, Newborn; Aquaporin 4; Astrocytes; Brain Edema; Erythropoietin; Glucose; Microscopy, Electron; Neuroprotective Agents; Oxygen; Phosphorylation; Primary Cell Culture; Protein Kinase C; Rats; Rats, Sprague-Dawley; Reperfusion Injury

There is a plethora of experimental data on the potential therapeutic benefits of recombinant human erythropoietin (rhEPO) and its synthetic derivatives in critical care medicine, in particular in ischemia/reperfusion injury. Most of the recent clinical trials have not shown clear benefits, and, in some patients, EPO-aggravated morbidity and mortality was even reported. Treatment with rhEPO has been successfully used in patients with anemia resulting from chronic kidney disease, but even a subset of this patient population does not adequately respond to rhEPO therapy. The following viewpoint uses rhEPO as an example to highlight the possible pitfalls in current practice using young healthy animals for the evaluation of therapies to treat patients of variable age and underlying chronic co-morbidity.

Topics: Acute Kidney Injury; Animals; Clinical Trials as Topic; Critical Illness; Drug Resistance; Erythropoietin; Humans; Models, Animal; Recombinant Proteins; Renal Insufficiency, Chronic; Reperfusion Injury

The objective of this study was to investigate the effects of erythropoietin (EPO) on systemic and renal hemodynamics in a rat model of renal ischemic/reperfusion (I/R) injury. We used 30 male Sprague-Dawley rats distributed among the following 3 groups (10 rats per group): (i) the sham-operated group, (ii) the control group (I/R injury only), and (iii) the EPO-treated group (I/R injury with 1500 U EPO·(kg body mass)⁻¹ on day 0, and 500 U·kg⁻¹ on days 2 and 4 after ischemia). Renal function, arterial blood pressure (ABP), renal plasma flow (RPF), renal blood flow (RBF), and renal vascular resistance (RVR) were measured on days 1, 2, and 7 after ischemia. The expression of endothelial NO synthase (eNOS) and histopathology of kidney were evaluated on day 7. The contractility of aortic strips was recorded from the different groups. The results show that renal function and histopathology were significantly improved after treatment with EPO. Compared with the control group, the EPO-treated group showed a significant increase in RPF, RBF, haematocrite, ABP, eNOS expression, and a decrease in RVR (p < 0.05).The response of aortic strips to the relaxant effect of acetylcholine was improved in the EPO-treated group. In conclusion, treatment with EPO improves renal function and renal haemodynamics in renal I/R injury, and causes significant rise of ABP and haematocrite value.

Topics: Animals; Aorta, Thoracic; Drug Resistance; Enzyme Induction; Epoetin Alfa; Erythropoietin; Hematinics; Hemodynamics; Hypotension; Ischemia; Kidney; Male; Nitric Oxide Donors; Nitric Oxide Synthase Type III; Protective Agents; Random Allocation; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Renal Circulation; Reperfusion Injury; Vascular Resistance; Vasodilation; Vasodilator Agents

New therapeutic strategies for chronic kidney disease (CKD) are necessary to offset the rising incidence of CKD and donor shortage. Erythropoietin (EPO), a cytokine produced by fibroblast-like cells in the kidney, has recently emerged as a renoprotective factor with anti-inflammatory, antioxidant properties. This study (a) determined whether human renal cultures (human primary kidney cells [hPKC]) can be enriched in EPO-positive cells (hPKC(F+)) by using magnetic-bead sorting; (b) characterized hPKC(F+) following cell separation; and (c) established that intrarenal delivery of enriched hPKC(F+) cells would be more beneficial in treatment of renal injury, inflammation, and oxidative stress than unsorted hPKC cultures in a chronic kidney injury model. Fluorescence-activated cell sorting analysis revealed higher expression of EPO (36%) and CD73 (27%) in hPKC(F+) as compared with hPKC. After induction of renal injury, intrarenal delivery of hPKC(F+) or hPKC significantly reduced serum creatinine, interstitial fibrosis in the medulla, and abundance of CD68-positive cells in the cortex and medulla (p < .05). However, only hPKC(F+) attenuated interstitial fibrosis in the renal cortex and decreased urinary albumin (3.5-fold) and urinary tubular injury marker kidney injury molecule 1 (16-fold). hPKC(F+) also significantly reduced levels of renal cortical monocyte chemotactic protein 1 (1.8-fold) and oxidative DNA marker 8-hydroxy-deoxyguanosine (8-OHdG) (2.4-fold). After 12 weeks, we detected few injected cells, which were localized mostly to the cortical interstitium. Although cell therapy with either hPKC(F+) or hPKC improved renal function, the hPKC(F+) subpopulation provides greater renoprotection, perhaps through attenuation of inflammation and oxidative stress. We conclude that hPKC(F+) may be used as components of cell-based therapies for degenerative kidney diseases.

Topics: Animals; Blotting, Western; Cell Proliferation; Cell Separation; Cell- and Tissue-Based Therapy; Erythropoietin; Female; Humans; Inflammation; Kidney; Kidney Failure, Chronic; Male; Middle Aged; Oxidative Stress; Rats; Rats, Nude; Real-Time Polymerase Chain Reaction; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

To observe the permeability of recombinant human erythropoietin through placenta barrier and fetal blood-brain barrier after transient uteroplacental ischemia.. Rats on days 19 of pregnancy were divided into rhEPO treated group, ischemia-reperfusion group and sham operated group. Fetal ischemia in rhEPO treated group and ischemia-reperfusion group was induced by bilateral occlusion of the utero-ovarian artery for 20 minutes. Different dosage of 125I-rhEPO (2500 U/kg, 5000 U/kg, 7500 U/kg) was injected into the rats through caudal veins 30 min before injury in rhEPO treated group and sham-operated group. Saline was administered intravenously 30 min before the induction of hypoxic-ischemic injury in ischemia-reperfusion group. The amniotic fluid, placenta and fetal organs including brain, liver, heart, lung and kidney were collected to measure the radioactivity at 24h after injury.. 125I-rhEPO was detected in amniotic fluid, placenta and fetal organs. The radioactivity of 125I-rhEPO in these tissues increased gradually with the increased dose injected in rhEPO treated group and sham-operated group. There were significant differences in the radioactivity of 125I-rhEPO between rhEPO treated group and sham-operated group (P < 0.05).. The permeability of rhEPO through placental barrier and blood-brain barrier increased under the condition of fetal ischemia and hypoxia.

Topics: Animals; Blood-Brain Barrier; Erythropoietin; Female; Fetal Hypoxia; Maternal-Fetal Exchange; Permeability; Pregnancy; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury

To investigate effects of combination of erythropoietin (EPO) and epidermal growth factor (EGF) on renal ischaemia and on reactive oxygen species in a rat model.. In all, 90 male Sprague-Dawley rats were allocated into five groups of 18, designated: Sham; treated with right nephrectomy only; Control, subjected to left renal ischaemia for 45 min with no treatment; EPO-treated, as the control but with EPO pretreatment; EGF-treated, as the control but with EGF pretreatment; EPO + EGF-treated, as the control but with EPO and EGF pretreatment. Renal function, histopathology and malondialdehyde (MDA), superoxide dismutase (SOD) and reduced glutathione (GSH) levels in kidneys were assessed at 1, 2 and 7 days after ischaemia.. All rats except the controls had a significant improvement in serum creatinine, creatinine clearance and fractional excretion of Na(+) ; all three were significantly better in EPO + EGF group than in all other groups Histopathological examination showed marked structural damage in control rats. The tubular damage was least in the EPO + EGF group. The control group had a significant increase in MDA level and a significant decrease in SOD and GSH, while the EPO + EGF group had a marked significant reduction in MDA and increase in GSH and SOD.. The protection against ischaemia/reperfusion injury might be maximal when EPO and EGF are administered concomitantly, and their protective effect might be partly due to their antioxidant effects.

Topics: Analysis of Variance; Animals; Antioxidants; Drug Therapy, Combination; Epidermal Growth Factor; Erythropoietin; Kidney; Male; Random Allocation; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Oxidative stress-related renal pathologies apparently include rhabdomyolysis and ischemia/reperfusion phenomenon. These two pathologies were chosen for study in order to develop a proper strategy for protection of the kidney. Mitochondria were found to be a key player in these pathologies, being both the source and the target for excessive production of reactive oxygen species (ROS). A mitochondria-targeted compound which is a conjugate of a positively charged rhodamine molecule with plastoquinone (SkQR1) was found to rescue the kidney from the deleterious effect of both pathologies. Intraperitoneal injection of SkQR1 before the onset of pathology not only normalized the level of ROS and lipid peroxidized products in kidney mitochondria but also decreased the level of cytochrome c in the blood, restored normal renal excretory function and significantly lowered mortality among animals having a single kidney exposed to ischemia/reperfusion. The SkQR1-derivative missing plastoquinone (C12R1) possessed some, although limited nephroprotective properties and enhanced animal survival after ischemia/reperfusion. SkQR1 was found to induce some elements of nephroprotective pathways providing ischemic tolerance such as an increase in erythropoietin levels and phosphorylation of glycogen synthase kinase 3β in the kidney. SkQR1 also normalized renal erythropoietin level lowered after kidney ischemia/reperfusion and injection of a well-known nephrotoxic agent gentamicin.

Topics: Animals; Antioxidants; Blotting, Western; Cells, Cultured; Erythropoietin; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Kidney; Kidney Diseases; Male; Microscopy, Confocal; Mitochondria; Molecular Structure; Oxidative Stress; Phosphorylation; Plastoquinone; Protective Agents; Rats; Reperfusion Injury; Rhabdomyolysis; Rhodamines; Survival Rate

The renoprotective effect of erythropoietin (Epo) against ischaemia-reperfusion injury (IR/I) was evaluated in a non-human primate model.. Crab-eating macaques were divided into two groups: Control (n = 10), treated with saline, and EPO (n = 10), treated with Epo. Epo was injected intravenously at a dose of 12,000 units, 5 min before clamping the renal pedicle and 5 min before declamping. Renal IR/I was created by clamping the left renal artery for 90 min following right nephrectomy. Haemoglobin (Hb), haematocrit (Ht), creatinine (Cr), blood urea nitrogen (BUN), cystatin C and interleukin-6 (IL-6) were measured before (Pre) and after (Day 0) the operation, and on post-operative days: Day 1, Day 3, Day 5 and Day 7. Apoptotic cells were counted on Day 1.. There were no differences in Hb and Ht between the two groups. Cr, BUN, cystatin C and IL-6 levels in the EPO group were lower than those in the Control group at most of the observation points. The number of apoptotic cells in the Control was significantly higher than that of and EPO group.. Epo significantly ameliorates renal IR/I in this non-human primate model. Our findings justify the clinical application of Epo, not only for acute renal failure, but also in transplantation.

Topics: Animals; Apoptosis; Creatinine; Disease Models, Animal; Erythropoietin; Humans; Kidney Function Tests; Macaca fascicularis; Male; Monkey Diseases; Reperfusion Injury

Paraplegia remains a devastating complication for patients undergoing thoracic aortic procedures. Although surgical adjuncts have evolved to reduce the risk of paraplegia, no pharmacologic therapies have proven efficacious in attenuating spinal cord ischemia-reperfusion injury. Effects of erythropoietin in spinal cord ischemia-reperfusion injury, however, have not yet been elucidated. We hypothesized that pretreatment with erythropoietin would attenuate functional and cytoarchitectural spinal cord injury related to high-risk aortic procedures.. Adult male mice were subjected to ischemia-reperfusion. Aortic arch and proximal left subclavian arteries were clamped for 5 minutes; animals were observed for 48 hours. Neurologic scores of hind limb function were assessed every 12 hours. Experimental groups consisted of treatment with erythropoietin 4 hours before crossclamping (n = 7), ischemic controls (n = 7), and sham ischemia (operation without crossclamping, n = 6). Thoracolumbar sections of spinal cord were removed after 48 hours and preserved for cytoarchitectural analysis.. Mice pretreated with erythropoietin exhibited significant preservation of hind limb motor function. All mice without pretreatment were paralyzed at 48 hours. Mice with erythropoietin pretreatment had improved motor function; 3 had no measurable neurologic deficit at 48 hours. Histologic analysis in mice treated with erythropoietin showed markedly reduced neuronal cell injury.. Erythropoeitin preserves both function and histologic appearance in mice undergoing spinal cord ischemia-reperfusion. With further elucidation of mechanisms of protection and optimal administration, erythropoietin could become an important adjunct in reducing the incidence and severity of spinal cord injury related to aortic interventions.

Topics: Animals; Aorta, Thoracic; Constriction; Disease Models, Animal; Erythropoietin; Hindlimb; Male; Mice; Mice, Inbred C57BL; Motor Activity; Muscle, Skeletal; Neuroprotective Agents; Paraplegia; Recombinant Proteins; Reperfusion Injury; Spinal Cord; Spinal Cord Ischemia; Subclavian Artery; Time Factors

Erythropoietin (EPO) is a cytokine with organ-protective properties. We hypothesized that EPO could attenuate acute renal dysfunction and inflammation in a porcine model of ischemia-reperfusion (IR). Furthermore, we aimed to characterize the impact of EPO on systemic and renal hemodynamics, and renal oxygen consumption.. Twenty-four pigs were randomly assigned to three groups: (1) EPO (5000 IU/kg) administered intravenously before IR (n=9), (2) placebo administered before IR (n=9), or (3) sham group, anesthetized and operated on only (n=6). IR was induced by clamping the left renal artery for 45 min. Hemodynamics and renal blood flow (RBF) were analyzed continuously. Glomerular filtration rate (GFR), renal oxygen consumption, and plasma cytokines (IL-1β, IL-6, IL-8, IL-10, and TNF-α) were analyzed hourly. Renal biopsies were analyzed for cytokine content and apoptosis.. GFR was higher during reperfusion in the EPO group than in the placebo group (P<0.01). No differences between the IR groups were found in hemodynamics, RBF, oxygen consumption, or renal apoptosis. The levels of TNF-α in the plasma (P=0.036) and the levels of TNF-α and IL-10 in the renal cortex (P=0.04 and P=0.01, respectively) were lower in the EPO group compared with the sham group.. EPO attenuated the renal dysfunction as estimated as GFR. This effect was not related to changes in the hemodynamics. The immunomodulatory effects of EPO were manifested as decreased levels of TNF-α and IL-10 in renal biopsies and TNF-α levels in plasma.

Topics: Acute Kidney Injury; Animals; Apoptosis; Caspase 3; Cytokines; Erythropoietin; Glomerular Filtration Rate; Hemodynamics; Immunohistochemistry; Kidney; Kidney Function Tests; Oxygen Consumption; Pulmonary Gas Exchange; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Renal Circulation; Reperfusion Injury; Swine

Ligustilide, the main lipophilic component of Danggui, has been reported to protect the brain against ischaemic injury. However, the mechanisms are unknown. Here, we investigated the roles of erythropoietin (EPO) and the stress-induced protein RTP801 in neuroprotection provided by ligustilide against ischaemia-reperfusion (I/R) damage to the brain.. The efficacy of ligustilide against I/R damage was assessed by neurological deficit, infarct volume and cell viability, using the middle cerebral artery occlusion model in rats in vivo and rat cultured neurons in vitro. EPO and RTP801 were analysed by Western blot. Over-expression of RTP801 was achieved by transfection of an expression plasmid.. Ligustilide decreased the neurological deficit score, infarct volume and RTP801 expression and increased EPO transcription in I/R rats, and increased cell viability and EPO and decreased LDH release and RTP801 in I/R neurons. Also, ligustilide increased ERK phosphorylation (p-ERK). The positive effects of ligustilide on p-ERK, cell viability and EPO were blocked by PD98059, but not LY294002 and SB203580. In addition, transfection of SH-SY5Y cells with RTP801 plasmid increased RTP801 and LDH release, while ligustilide inhibited the effects of transfection on RTP801 expression and also increased cell viability.. Ligustilide exerts neuroprotective effects against I/R injury by promoting EPO transcription via an ERK signalling pathway and inhibiting RTP801 expression, This compound could be developed into a therapeutic agent to prevent and treat ischaemic disorders.

Topics: 4-Butyrolactone; Animals; Cell Line; Chromones; Erythropoietin; Flavonoids; Gene Expression Regulation; Glucose; Imidazoles; L-Lactate Dehydrogenase; Male; Morpholines; Neurons; Oxygen; Plasmids; Pyridines; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Repressor Proteins; Transcription Factors

Perioperative renal dysfunction is associated with a high mortality. The aim of this study was to investigate whether isoflurane preconditioning provides a protection against renal ischemic-reperfusion injury and whether hypoxia inducible factor 1 α (HIF-1 α) is responsible for the protection afforded by isoflurane in mice.. Adult male C57BL/6 mice received vehicle (PBS), scrambled siRNA or HIF-1 α siRNA via hydrodynamic injection through tail vein. Twenty-four hours after injection, they were exposed to 1.5% isoflurane in oxygen enriched air for 2h while controls without injection were exposed to oxygen enriched air. Twenty-four hours after gas exposure, mice were sacrificed and their kidney were harvested for western blot while other cohorts underwent renal ischemia-reperfusion injury induced by bilateral renal pedicle clamping for 25 min for renal histological or functional analysis 24h after reperfusion or by unilateral clamping for 40 min for survival rate analysis.. Survival rate and the expression of HIF-1 α and erythropoietin were significantly increased while apoptosis, renal tubule score, blood plasma creatinine and urea were decreased by isoflurane preconditioning. HIF-1 α siRNA but not scrambled siRNA injection abrogated the protective effect of isoflurane preconditioning.. Our data suggested that isoflurane preconditioning provided a protection against renal ischemic-reperfusion injury which is very likely due to hypoxia inducible factor-1 α upregulation.

Topics: Animals; Apoptosis; Blotting, Western; Creatinine; Disease Models, Animal; Erythropoietin; Hypoxia-Inducible Factor 1, alpha Subunit; In Situ Nick-End Labeling; Isoflurane; Kidney; Male; Mice; Mice, Inbred C57BL; Protective Agents; Reperfusion Injury; RNA, Small Interfering; Time Factors; Urea

Renal ischemia reperfusion injury induces gender-dependent heat-shock protein 72 expression, which maintains membrane localization of renal Na(+)/K(+)ATPase-α1. The erythropoietin has a protecting effect against ischemia reperfusion injury in various organs. In this study, we investigated whether erythropoietin exerts a beneficial effect against post-ischemic renal injury. Furthermore, we studied the erythropoietin signaling on heat-shock protein 72 and Na(+)/K(+)ATPase-α1 expression and localization.. In male and female Wistar rats, rHuEPO (1000 IU/bwkg intraperitoneal) or vehicle was administered 24 hours prior to unilateral left renal ischemia reperfusion (50 minutes). Kidneys were subsequently removed at hours 2 or 24 of the reperfusion; sham-operated rats served as controls (C) (n = 8/group). We measured serum erythropoietin, renal function, evaluated histological injury, and observed heat-shock protein 72 as well as Na(+)/K(+)ATPase-α1 protein level and localization. Additional groups were followed for 7-day survival.. Erythropoietin treatment was associated with better post-ischemic survival and less impaired renal function in males while diminishing the renal structural damage in both sexes. Endogenous erythropoietin was higher in males and increased in both genders after erythropoietin treatment. The erythropoietin treatment elevated protein levels of heat-shock protein 72 and Na(+)/K(+)ATPase-α1 in 24 hours in males, whereas in females, the already higher expression of heat-shock protein 72 and Na(+)/K(+)ATPase-α1 was not increased. Moreover, erythropoietin prevented ischemia reperfusion induced Na(+)/K(+)ATPase-α1 translocation from the basolaterale membrane in males.. Erythropoietin diminishes gender difference in the susceptibility to renal post-ischemic injury and reduces post-ischemic structural damage while preserving kidney function, particularly in males. This additional protection may be associated with a heat-shock protein 72-mediated effect on Na(+)/K(+)ATPase-α1 expression and translocation.

Topics: Animals; Blood Urea Nitrogen; Creatinine; Erythropoietin; Female; HSP72 Heat-Shock Proteins; Immunohistochemistry; Kidney; Male; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury; Sex Characteristics; Signal Transduction; Sodium-Potassium-Exchanging ATPase

The neuroprotective effect of erythropoietin has been demonstrated by ischemia and reperfusion models in adult and neonatal rodents. However, administration of high-dose erythropoietin has potential complications. The goal of this study was to determine whether local infusion of low dose erythropoietin offers neuroprotective effects after ischemia and reperfusion injury.. Adult male Sprague-Dawley rats subject to middle cerebral artery occlusion were randomly divided into three groups: (1) sham group: the rats received the same procedure as the other two groups except that no suture was inserted; (2) vehicle group: intra-artery local infusion of saline was administered via middle cerebral artery after reperfusion; and (3) treatment group: 50 U/kg intra-artery local infusion of erythropoietin was administered via middle cerebral artery after reperfusion. Neurological deficit scores and infarct volume (determined by hematoxylin-eosin staining) were evaluated 48 hours after reperfusion. Apoptosis was measured through terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. The expression of vascular endothelial growth factor and phosphorylated extracellular signal-regulated kinase were investigated by immunohistochemistry method.. The results show that intra-artery local infusion of erythropoietin, via the middle cerebral artery, significantly reduced neurological deficit scores, foot fault number, and the infarct volume at 48 hours after reperfusion. Significant reductions were also found in the number of positive cells stained by TUNEL assay within the ischemic core and penumbra. Furthermore, local infusion of erythropoietin increased the expression of phosphorylated extracellular signal-regulated kinase and vascular endothelial growth factor.. Local infusion of low-dose erythropoietin via the middle cerebral artery is shown to be neuroprotective against cerebral ischemia and reperfusion injury. The mechanism of neuroprotection may be associated with the increased expression of phosphorylated extracellular signal-regulated kinase and vascular endothelial growth factor.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Erythropoietin; Infarction, Middle Cerebral Artery; Infusions, Intra-Arterial; Male; Neuroprotective Agents; Random Allocation; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Ischemia/reperfusion (I/R) injury is an important cause of renal graft dysfunction. Increases in cold and warm ischemia times lead to a higher risk of early posttransplant complications including delayed graft function and acute rejection. Moreover, prolonged cold ischemia is a predictor of long-term graft loss in kidney transplant patients.. Darbepoetin alfa (DA) and carbamylated nonerythropoietic derivative of erythropoietin (CEPO) protective effects were evaluated in a model of I/R injury after kidney transplantation in both syngeneic and allogeneic combinations. The effects of wortmannin (phosphorylated Akt [p-Akt] inhibitor) administration were also investigated. Serum creatinine was evaluated at 16, 24, 48 hr and at 4 and 7 days posttransplant. Animals were killed 24 hr or 7 days after transplant and kidneys were processed for histological analysis, immunohistochemistry assessment of erythropoietin receptor (EPOR) and β-common chain receptor expression, granulocyte infiltration, nitrotyrosine staining, p-Akt expression, peritubular capillary (PTC) density, apoptosis, antioxidant, and antiapoptotic gene expression.. DA and CEPO significantly reduced serum creatinine, tubular injury, tubular nitrotyrosine staining, and prevented I/R-induced tubular apoptosis, but only when given both to the donor and to the recipient. DA and CEPO cytoprotection was associated with prevention of I/R-induced drop of p-Akt expression in tubuli, and almost complete preservation of capillary density in the tubulointerstitium of the graft. CEPO was more effective than DA in reducing tubular oxidative stress and preserving PTCs.. DA and CEPO when given both to the donor and to the recipient, prevented renal graft dysfunction, tubular oxidative stress, and apoptosis after I/R injury in kidney transplantation. Their cytoprotection was mediated by tubular p-Akt activation and PTC density preservation.

Topics: Androstadienes; Animals; Antioxidants; Apoptosis; Apoptosis Regulatory Proteins; Cold Temperature; Darbepoetin alfa; Erythropoietin; Graft Survival; Hematinics; Immunosuppressive Agents; Kidney Transplantation; Kidney Tubules; Oxidative Stress; Primary Graft Dysfunction; Proto-Oncogene Proteins c-akt; Rats; Rats, Inbred Lew; Reperfusion Injury; Tissue Donors; Transplantation, Homologous; Wortmannin

Topics: Animals; Aorta; Catheterization; Erythropoietin; Reperfusion Injury; Spinal Cord Ischemia

Recombinant human erythropoietin (rhEPO) attenuated ischemia/reperfusion (I/R) injury-induced spinal cord damage. Since carbamylated EPO derivatives are stated to be devoid of rhEPO side effects, we tested the hypothesis that a newly developed carbamylated EPO-FC fusion protein (cEPO-FC) would compare favorably with rhEPO.. Anesthetized and mechanically ventilated pigs randomly received cEPO-FC (50 μg kg(-1)), rhEPO (5,000 IU kg(-1)) or vehicle (n = 9 per group) 30 min prior to 30 min of aortic occlusion and over the 4 h of reperfusion. During aortic occlusion, mean arterial pressure (MAP) was maintained at 80-120% of baseline values by esmolol, nitroglycerin, and adenosine-5'-triphosphate (ATP). During reperfusion, noradrenaline was titrated to keep MAP at pre-ischemic levels. Spinal cord function was assessed by motor evoked potentials (MEP) and lower limb reflexes. Tissue damage was evaluated using hematoxylin and eosin, Nissl, and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining. Plasma levels of interleukin-6, tumor necrosis factor-α, and 8-isoprostanes were measured as markers of systemic inflammation and oxidative stress.. While only cEPO-FC restored MEP amplitude to values close to pre-occlusion levels, both cEPO-FC and rhEPO comparably restored lower limb reflexes and reduced the percentage of damaged neurons. Infiltration of mononuclear inflammatory cells was moderate without intergroup difference; positive TUNEL staining was barely detectable in any group. I/R injury increased blood cytokine levels without intergroup difference, whereas both cEPO-FC and rhEPO significantly lowered 8-isoprostane levels.. In a porcine model of aortic balloon occlusion-induced spinal cord I/R injury, cEPO-FC and rhEPO comparably protected against ischemic spinal cord dysfunction and neuronal damage. This effect coincided with attenuated oxidative stress.

Topics: Animals; Aorta; Catheterization; Erythropoietin; Evoked Potentials, Motor; Reperfusion Injury; Spinal Cord Ischemia; Swine

Topics: Animals; Disease Models, Animal; Erythropoietin; Humans; Male; Monkey Diseases; Reperfusion Injury

Prolyl hydroxylases (PHDs) are oxygen sensors that stabilize hypoxia-inducible factors (HIFs) to induce proinflammatory, vasopermeability, and proapoptotic factors. These may be potential targets to reduce the complications of ischemic retinopathies.. Oxygen-induced ischemic retinopathy (OIR) was generated as a model for retinopathy of prematurity (ROP) by placing 7-day-old mice in 75% oxygen for 5 days and returning them to the relative hypoxia of room air for 5 days. Neovascularization (NV) and avascular areas were assessed on retinal flat-mounts by image analysis. Blood-retinal barrier breakdown was assessed using ³H-mannitol as a tracer. Apoptosis was detected with TUNEL staining. HIF-1α and VEGF were quantified using Western blot analysis and ELISA.. PHD1-deficient mice demonstrated reduced hyperoxia-associated vascular obliteration during oxygen-induced ischemic retinopathy. This was associated with subsequent reduced avascularity, vascular leakage, and pathologic NV during the hypoxic phase, which could be accounted for by a reduced expression of HIF-1α and VEGF. Apoptosis in the retina was also reduced in PHD1-depleted mice after 2 days in hyperoxia.. PHD1 deficiency is associated with a reduction of ischemia-induced retinal NV. The regulatory mechanism in this model appears to be: PHD1 depletion prevents HIF-1α degradation in hyperoxia, which induces VEGF, thus preventing hyperoxia-related vessel loss. Without a vessel deficiency, there would not be relative hypoxia when the mice are returned to room air and there would be no need to initiate angiogenesis signaling. Blocking PHD1 may be beneficial for ischemic retinopathies and inflammatory and neurodegenerative disorders.

Topics: Animals; Animals, Newborn; Apoptosis; Blood-Retinal Barrier; Blotting, Western; Capillary Permeability; Enzyme-Linked Immunosorbent Assay; Erythropoietin; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; In Situ Nick-End Labeling; Infant, Newborn; Mice; Mice, Knockout; Oxygen; Procollagen-Proline Dioxygenase; Reperfusion Injury; Retinal Neovascularization; Retinal Vessels; Retinopathy of Prematurity; Vascular Endothelial Growth Factor A

Recent research based on various animal models has shown the neuroprotective effects of erythropoietin (EPO). However, few studies have examined such effects of EPO in the clinic. In this study we enrolled patients with spinal cord ischemia-reperfusion (I-R) injury to investigate the clinical application of EPO and methylprednisolone (MP) for the neuroprotection against spinal cord I-R injury. Retrospective analysis of 63 cases of spinal cord I-R injury was performed. The Frankel neurological performance scale was used to evaluate the neurological function after spinal cord injury (SCI), including 12 cases of scale B, 30 cases of scale C, and 21 cases of scale D. These cases were divided into 2 groups: group A (27 cases) got treatment with both EPO and MP; group B (36 cases) got treatment with MP only. The neurological function of patients after treatment was evaluated by American Spinal Cord Injury Association (ASIA) index score, and activity of daily living (ADL) of the patients was also recorded. All patients got follow-up and the follow-up period ranged from 24 to 39 months (mean 26 months). There was no significance difference in neurological function between groups A and B before the treatment (P>0.05). However, the neurological function and ADL scores were significantly improved 1 week, 1 year or 2 years after the treatment compared to those before the treatment (P<0.05), and the improvement was more significant in group A than in group B (P<0.05). It is suggested that the clinical application of EPO and MP provides the neuroprotection against spinal cord I-R injury.

Topics: Adult; Aged; Decompression, Surgical; Erythropoietin; Female; Humans; Male; Methylprednisolone; Middle Aged; Neuroprotective Agents; Reperfusion Injury; Retrospective Studies; Spinal Cord; Spinal Cord Compression; Spinal Cord Ischemia

Hyperglycemia is associated with a decreased tolerance to ischemia and an increased severity of renal ischemia reperfusion (I/R) injury. It has been suggested that erythropoietin (EPO) attenuates this effect in normoglycemic animals. This study sought to examine the effects of EPO on treatment renal I/R injury (IRI) in transiently hyperglycemic rats.. Twenty-eight male Wister rats anesthetized with isoflurane received glucose (2.5 g.kg(-1) intraperitoneally) before right nephrectomy. They were randomly assigned to four groups: sham operation (S); IRI (ISO); IRI+EPO, (600 UI kg(-1) low-dose EPO [EL]); and IRI+EPO 5000 UI kg(-1) (high-dose EPO [EH]). IRI was induced by a 25-minute period of left renal ischemia followed by reperfusion for 24 hours. Serum creatinine and glucose levels were measure at baseline (M1), immediately after the ischemic period (M2), and at 24 hours after reperfusion (M3). After sacrificing the animals, left kidney specimens were submitted for histological analysis including flow cytometry to estimate tubular necrosis and the percentages of apoptotic, dead or intact cells.. Scr in the ISO group was significantly higher at M3 than among the other groups. Percentages of early apoptotic cells in ISO group were significantly higher than the other groups. Percentages of late apoptotic cells in S and ISO groups were significantly greater than EL and EH groups. However, no significant intergroup differences were observed regarding the incidence of tubular necrosis.. Our results suggested that, although not preventing the occurrence of tubular necrosis, EPO attenuated apoptosis and glomerular functional impairment among transiently hyperglycemic rats undergoing an ischemia/reperfusion insult.

Topics: Animals; Apoptosis; Blood Glucose; Creatinine; Cytoprotection; Disease Models, Animal; Epoetin Alfa; Erythropoietin; Flow Cytometry; Hyperglycemia; Kidney; Kidney Glomerulus; Kidney Tubules; Male; Necrosis; Nephrectomy; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury; Time Factors

To increase the survival rate of patients with acute superior mesenteric artery thromboembolism (ASMAT) treated by catheter thrombolysis, we examined the effects of delivering edaravone and asialoerythropoietin, agents with tissue-protective activities, using a rabbit autologous fibrin clot ASMAT model. Japanese white rabbits (n=32) were randomly separated into four equal groups. 45 min after introducing autologous fibrin clot, Group U received urokinase and heparin; Group E received urokinase and heparin plus edaravone; Group A received urokinase and heparin plus asialoerythropoietin; and Group EA received urokinase, heparin and edaravone plus asialoerythropoietin via a catheter. The intestines were removed 6 h later and intestinal mucosal damage was scored using the Park's injury score. Survival time was assessed. Average mucosal injury was 5.78+/-1.52 (Group U), 2.88+/-0.72 (Group E), 1.90+/-1.23 (Group A) and 1.18+/-1.25 (Group EA). The degree of mucosal injury was significantly lower in Group EA than in Groups U and E (p<0.05). Conversely, there was no significant difference between Group A and Group EA, or between Group A and Group E. The survival times were 31.50+/-13.30 h (Group U), 51.00+/-24.74 h (Group E), 48.00+/-16.97 h (Group A) and 82+/-51.07 h (Group EA); the difference among the four groups was not significant. In conclusion, the concomitant administration of asialoerythropoietin and edaravone reduced mucosal membrane injury significantly compared with edaravone alone. However, to improve the survival of ASMAT rabbit models, the delivery of an appropriate dose of asialoerythropoietin is required, together with the development of methods to assess peripheral recanalisation.

Topics: Animals; Antipyrine; Asialoglycoproteins; Catheterization; Disease Models, Animal; Drug Combinations; Edaravone; Erythropoietin; Fibrin; Fibrinolytic Agents; Free Radical Scavengers; Heparin; Injections, Intra-Arterial; Intestinal Mucosa; Mesenteric Artery, Superior; Mesenteric Vascular Occlusion; Rabbits; Random Allocation; Reperfusion Injury; Survival Rate; Thromboembolism; Urokinase-Type Plasminogen Activator

Erythropoietin (EPO) protects the kidneys from ischemia/reperfusion (I/R) injury; however, the exact signalling mechanisms are not fully understood. The serum and glucocorticoid-regulated kinase 1 (SGK1) is an anti-apoptotic protein kinase regulated through the phosphatidylinositol 3-kinase (PI3-kinase) pathway by cellular stimuli, hormones and growth factors. The objective of the present study was to examine the role of SGK1 in the renoprotective effects of EPO in renal I/R injury. In vitro, cultures of HEK293 cells were exposed to 16h hypoxia. Incubation with EPO at a doses of 400U/ml exerted a protective effect on cell death assessed by LDH release and Annexin V FACS analysis. This was paralleled by up-regulation of SGK1 expression, as well as phosphorylation. Downregulation of SGK1 expression by small interfering RNA technique ameliorated the anti-apoptotic effect of EPO treatment. In an in vivo rat model of unilateral renal I/R injury, rats were treated with 500U/kg EPO 24h prior to ischemia. EPO resulted in less severe tissue injury and ameliorated the elevation in creatinine and urea nitrogen levels 24h after reperfusion. Furthermore, SGK1 expression and phosphorylation were higher in EPO compared to vehicle-treated rats as demonstrated by real-time PCR, Western blot and immunofluorescence technique. We conclude that EPO protects from renal I/R injury and SGK1 might contribute to the mediation of EPO effects under ischemic conditions.

Topics: Animals; Apoptosis; Cells, Cultured; Erythropoietin; Humans; Immediate-Early Proteins; Kidney; Male; Phosphorylation; Protein Serine-Threonine Kinases; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury

A noninvasive assessment of infarct size and transmural extension of myocardial infarction (TEMI) is fundamental in experimental models of ischemia-reperfusion. Conventional echocardiography parameters are limited in this purpose. This study was designed to examine whether speckle tracking imaging can be used in a rat model of ischemia-reperfusion to accurately detect the reduction of infarct size and TEMI induced by erythropoietin (EPO) as early as 24 h after reperfusion. Rats were randomly assigned to one of three groups: myocardial infarction (MI)-control group, 45 min ischemia followed by 24 h of reperfusion; MI-EPO group, similar surgery with a single bolus of EPO administered at the onset of reperfusion; and sham-operated group. Short-axis two-dimensional echocardiography was performed after reperfusion. Global radial (GS(r)) and circumferential (GS(cir)) strains were compared with infarct size and TEMI assessed after triphenyltetrazolium chloride staining. As a result, ejection fraction, shortening fraction, GS(r), and GS(cir) significantly correlated to infarct size, whereas only GS(r) and GS(cir) significantly correlated to TEMI. EPO significantly decreased infarct size (30.8 + or - 3.5 vs. 56.2 + or - 5.7% in MI-control, P < 0.001) and TEMI (0.37 + or - 0.05 vs. 0.77 + or - 0.05 in MI-control, P < 0.001). None of the conventional echocardiography parameters was significantly different between the MI-EPO and MI-control groups, whereas GS(r) was significantly higher in the MI-EPO group (29.1 + or - 4.7 vs. 16.4 + or - 3.3% in MI-control; P < 0.05). Furthermore, GS(cir) and GS(r) appeared to be the best parameters to identify a TEMI >0.75 24 h after reperfusion. In conclusion, these findings demonstrate the usefulness of speckle tracking imaging in the early evaluation of a cardioprotective strategy in a rat model of ischemia-reperfusion.

Topics: Animals; Echocardiography, Doppler; Erythropoietin; Models, Animal; Myocardial Infarction; Myocardium; Necrosis; Rats; Rats, Wistar; Reperfusion Injury; Salvage Therapy; Time Factors

Diabetic patients are at higher risk of failure to recover after acute kidney injury, however, the mechanism and therapeutic strategies remain unclear. Erythropoietin is cytoprotective in a variety of non-haematopoietic cells. The aim of the present study was to clarify the mechanism of diabetes-related acceleration of renal damage after ischaemia-reperfusion injury and to examine the therapeutic potential of asialoerythropoietin, a non-haematopoietic erythropoietin derivative, against ischaemia-reperfusion-induced acute kidney injury in diabetic mice.. C57BL/6J mice with and without streptozotocin-induced diabetes were subjected to 30 min unilateral renal ischaemia-reperfusion injury at 1 week after induction of diabetes. They were divided into four group: (i) non-diabetic plus ischaemia-reperfusion injury; (ii) non-diabetic plus ischaemia-reperfusion injury plus asialoerythropoietin (3000 IU/kg bodyweight); (iii) diabetic plus ischaemia-reperfusion injury; and (iv) diabetic plus ischemia-reperfusion injury plus asialoerythropoietin. Experiments were conducted at the indicated time periods after ischaemia-reperfusion injury.. Ischaemia-reperfusion injury of diabetic kidney resulted in significantly low protein expression levels of bcl-2, an anti-apoptotic molecule, and bone morphogenetic protein-7 (BMP-7), an anti-fibrotic and pro-regenerative factor, compared with non-diabetic kidneys. Diabetic kidney subsequently showed severe damage including increased tubular cell apoptosis, tubulointerstitial fibrosis and decreased tubular proliferation, compared with non-diabetic kidney. Treatment with asialoerythropoietin induced bcl-2 and BMP-7 expression in diabetic kidney and decreased tubular cell apoptosis, tubulointerstitial fibrosis and accelerated tubular proliferation.. Reduced induction bcl-2 and BMP-7 may play a role in the acceleration of renal damage after ischaemia-reperfusion injury in diabetic kidney. The renoprotective effects of asialoerythropoietin on acute kidney injury may be mediated through the induction of bcl-2 and BMP-7.

Topics: Acute Kidney Injury; Animals; Asialoglycoproteins; Diabetes Mellitus, Experimental; Erythropoietin; Male; Mice; Mice, Inbred C57BL; Reperfusion Injury

The only currently offered curative option for many patients with primary or secondary liver tumors is the resection of hepatic tumors. The aim of this study was to evaluate the role of recombinant human erythropoietin (rhEPO) in liver protection and regeneration after subtotal hepatectomy in rats. Rats undergoing 70% hepatectomy received an intraperitoneal injection of saline (control) or rhEPO (4 U/g) 30 minutes prior to resection. Liver function was assessed by the measurement of the international normalized ratio (INR) levels, and hepatic injury was assessed by serum alanine aminotransferase and aspartate aminotransferase levels. Hepatic apoptosis was assessed by intrahepatic caspase-3 activity and morphological criteria. The regeneration capacity of remnant livers was assessed over 7 days with the regenerated liver/body weight ratio, immunohistochemistry markers of cell proliferation (Ki-67) and angiogenesis (von Willebrand factor), and phosphorylated extracellular signal-regulated kinase signaling. Two and 4 days after subtotal hepatectomy, the regenerated liver/body weight ratio was significantly higher in animals treated with rhEPO versus the control group (P < 0.005). Serum liver enzymes and INR levels on days 2 and 4 post-hepatectomy were significantly lower in animals pretreated with rhEPO in comparison with the control group (P < 0.005). No statistically significant difference was noted in intrahepatic hepatic caspase-3 activity, immunohistochemistry for caspase-3, or a terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling assay between the hepatectomized groups. In the rhEPO-pretreated group, the mitotic index, Ki-67 and von Willebrand factor expression, and extracellular signal-regulated kinase activity were significantly higher on day 2 post-hepatectomy (P < 0.05) in comparison with the control group. In conclusion, rhEPO treatment may offer a unique beneficial dual-function strategy for hepatic protection and regeneration immediately after subtotal hepatectomy in rats.

Topics: Alanine Transaminase; Animals; Apoptosis; Aspartate Aminotransferases; Blotting, Western; Caspase 3; Erythropoietin; Hepatectomy; Humans; Immunohistochemistry; In Situ Nick-End Labeling; International Normalized Ratio; Ki-67 Antigen; Liver; Liver Regeneration; Male; Organ Size; Postoperative Complications; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury; von Willebrand Factor

While the problem of organ shortage has not yet been solved, the number of patients who need to be treated with dialysis due to end-stage renal disease (ESRD) is increasing each year. With the aim of eliminating dialytic therapy as much as possible, the opportunities for organ donation from expansive criteria donor (ECD) or marginal donors due to cardiac death have been increasing. With the purpose of extracting organs in a state in which the function is preserved as much as possible, we reexamined the conditions of tissue disorders resulting from temporary ischemia of the organs as well as changes in tissue function and the effects on the preservation of renal function over time by using rat models in order to clinically utilize erythropoietin, which has inhibitory effects on ischemia-reperfusion disorder, as has been conventionally reported. With 8- to 9-week-old Wister male rats, after the right kidney had been resected under general anesthesia, the left renal artery was clamped to inhibit the blood flow for 45 min. At 30 min before inhibiting the blood flow and after releasing the inhibited blood flow, 100 U/kg of recombinant human erythropoietin (rhEPO) was administered via the inferior vena cava and the abdominal cavity, and then the tissues and blood samples were extracted at 6 and 24 h after the release. The renal tissue specimens were evaluated using H&E staining and TUNEL staining in order to observe differences in the expression of apoptosis as well as the renal function and changes in the emergence of active oxygen were investigated by using samples that had been obtained from drawn blood. Moreover, we examined the degree of renal dysfunction by means of neutrophil gelatinase-associated lipocalin (NGAL) in the spot urine samples. The changes in renal function, which were observed according to the serum creatinine level, showed that the renal function was preserved with a significant difference in the rhEPO administration group. The liver deviation enzymes, which had also shown increases in the serum as well as the occurrence of renal dysfunction, showed clear decreases in the serum, even though changes with a significant difference were not observed in the rhEPO administration group. The active oxygen did not show changes before and after ischemia-reperfusion nor changes due to the rhEPO administration. When examining the status of apoptosis in the tissues, apoptosis was shown to be inhibited due to the rhEPO administration. It is believed tha

Topics: Acute-Phase Proteins; Animals; Blood Urea Nitrogen; Cell Count; Creatinine; Enzyme-Linked Immunosorbent Assay; Erythropoietin; Humans; In Situ Nick-End Labeling; Kidney; Lipocalin-2; Lipocalins; Liver; Male; Protective Agents; Proto-Oncogene Proteins; Rats; Reactive Oxygen Species; Recombinant Proteins; Reperfusion Injury

The aim of this study was to investigate the effects of erythropoietin and dimethylsulfoxide in the recovery from ischemia-reperfusion injury in an experimental rat adnexal torsion model.. Thirty-six Wistar-albino rats were divided into six groups. Except for the sham operation group, all groups were subjected to left unilateral adnexal torsion for 3h. Erythropoietin and dimethylsulfoxide were intraperitoneally administered 30min before the detorsion operation. Malondialdehyde and nitric oxide levels were detected from both the plasma and the tissue samples. The sections of the tissues were evaluated histologically. The results were analyzed by a one-way analysis of the variance (ANOVA) followed by the Duncan test for multiple comparisons using computer software, SPSS Version 15.0 for Windows.. This study demonstrated that dimethylsulfoxide and erythropoietin pretreatment attenuated ischemia-reperfusion-induced lipid peroxidation, prevented post-ischemic ovarian injury and helped to maintain the ovarian morphology. Malondialdehyde levels of plasma and ovary were higher in the torsion and detorsion groups than the sham group. This showed that ischemia-reperfusion had caused lipid peroxidation of the ovarian tissue, thus leading to oxidative damage. One of the major findings of this study is that malondialdehyde was significantly decreased in the plasma of rats who were pre-treated with dimethylsulfoxide and erythropoietin before detorsion. This suggests that dimethylsulfoxide and erythropoietin might prevent oxidative damage in ovarian ischemia-reperfusion injury. Histological examination confirmed that reperfusion caused more detrimental effects than only ischemia, which could be at least partially prevented by dimethylsulfoxide and erythropoietin administration prior to detorsion.. Erythropoietin and dimethylsulfoxide may have beneficial effects in ischemia-reperfusion injury in ovarian torsion.

Topics: Adnexal Diseases; Animals; Dimethyl Sulfoxide; Erythropoietin; Female; Malondialdehyde; Nitric Oxide; Ovarian Diseases; Ovary; Rats; Rats, Wistar; Reperfusion Injury; Torsion Abnormality

It has been demonstrated that hypoxia-inducible factor-1 alpha (HIF-1 alpha) mediates ischemic tolerance induced by hypoxia/ischemia or pharmacological preconditioning. In addition, preconditioning stimuli can be cross-tolerant, safeguarding against other types of injury. We therefore hypothesized HIF-1 alpha might also be associated with ischemic tolerance induced by hyperthermic preconditioning. In the present study, we demonstrated for the first time that 6 h of hyperthermia (38°C or 40°C) could induce a characteristic "reactive" morphology and a significant increase in the expression of bystin in astrocytes. We also showed that pre-treatment with 6 h of hyperthermia resulted in a significant increase in cell viability and a remarkable decrease in lactate dehydrogenase (LDH) release and apoptosis development in the astrocytes that were exposed to 24 h of ischemia and a subsequent 24 h of reperfusion. Analysis of mechanisms showed that hyperthermia could lead to a significant increase in HIF-1 alpha expression and also the HIF-1 binding activity in the ischemia/reperfusion astrocytes. The data provide evidence to our hypothesis that the up-regulation of HIF-1 alpha is associated with the protective effects of hyperthermic preconditioning on astrocytes against ischemia/reperfusion injury.

Topics: Animals; Animals, Newborn; Apoptosis; Astrocytes; Binding Sites; Blotting, Western; Cell Adhesion Molecules; Cell Shape; Cells, Cultured; Electrophoretic Mobility Shift Assay; Erythropoietin; Glial Fibrillary Acidic Protein; Hot Temperature; Hypoxia-Inducible Factor 1, alpha Subunit; Ischemic Preconditioning; L-Lactate Dehydrogenase; Mice; Mice, Inbred ICR; Nuclear Proteins; Protein Binding; Reperfusion Injury; Time Factors; Up-Regulation

Ischemia-reperfusion (I/R) injury may influence graft function after transplantation. Erythropoietin (EPO) attenuates I/R injury in various animal organs such as intestine, brain, and kidney.. To evaluate the effects of pretreatment with recombinant human EPO (rhEPO) on I/R-induced heart injury.. A rat model of I/R injury was established by ligating the left descending coronary artery for 30 minutes, followed by reperfusion for 4 hours. Fifty Sprague-Dawley rats were divided into 5 groups: sham operation; I/R; I/R+rhEPO, 100 U/kg; I/R+rhEPO, 1000 U/kg; and I/R+rhEPO, 5000 U/kg. Electrocardiograms were assessed continuously to note arrhythmia caused by reperfusion. Serum concentrations of interleukin (IL)-6 and IL-8, and tumor necrosis factor-alpha were measured at 2 and 4 hours after reperfusion.. The rhEPO-treated animals exhibited dosage-dependent significant reduction in the incidence of ventricular arrhythmia caused by reperfusion, and markedly decreased serum concentrations of IL-6, IL-8, and tumor necrosis factor-alpha (P < .05) compared with the I/R group (P < .05).. The rhEPO attenuates myocardial I/R injury in rats, at least in part related to inhibition of the system inflammatory response.

Topics: Animals; Erythropoietin; Humans; Inflammation; Interleukin-6; Interleukin-8; Myocardial Ischemia; Myocardial Reperfusion Injury; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Tumor Necrosis Factor-alpha

To investigate whether recombine human erythropoietin can cross the placenta barrier in rats with transient uteroplacental ischemia.. Rats on day 19 of pregnancy were divided into ischemia-reperfusion group, sham-operated group and rhEPO treated group. Fetal ischemia was induced by bilateral occlusion of the utero-ovarian artery for 20 minutes. 125I-rhEPO or saline were administered intravenously 30 min before the induction of hypoxic-ischemic injury. Fetal rat organs were removed to measure the radioactivity post injury.. A small amount of radioactive activity (1.26 +/- 0.28) pg/g was detected in the fetal rats with ischemia-reperfusion. 125I-rhEPO radioactivity increased gradually with time in the placenta, amniotic fluid and fetal tissues of vital organs in the rats of sham-operated group and rhEPO treatment group. There were significant differences in 125I-rhEPO between placental organs and other organs (P < 0.05). The permeability of 125I-rhEPO through the blood brain barrier changed with reperfusion time and peaked about 6 h after reperfusion, and significant differences were found between rhEPO treatment group and sham operation group (P < 0.05).. Exogenous rhEPO can cross the placenta barrier and blood-brain barrier and reach hypoxic-ischemic fetal rats.

Topics: Animals; Blood-Brain Barrier; Capillary Permeability; Erythropoietin; Female; Fetal Hypoxia; Ischemia; Maternal-Fetal Exchange; Placenta; Pregnancy; Rats; Recombinant Proteins; Reperfusion Injury; Uterus

Topics: Animals; Brain Edema; Brain Ischemia; Erythropoietin; Female; Hippocampus; Male; Nitric Oxide; Protective Agents; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Superoxide Dismutase

We reexamined the conditions of tissue disorders resulting from temporary ischemia of the organs as well as changes in tissue function and the effects on the preservation of renal function over time by using rat models in order to clinically utilize erythropoietin, which has inhibitory effects on ischemia-reperfusion disorders. In 8- to 9-week-old Wister male rats, after the right kidney had been resected under general anesthesia, the left renal artery was clamped to inhibit the blood flow for 45 minutes. At 30 minutes before inhibiting the blood flow and after releasing the inhibited blood flow, 100 U/kg of recombinant human erythropoietin (rhEPO) was administered via the inferior vena cava and the abdominal cavity, and then the tissues and blood samples were extracted at 6 hours and 24 hours after the release. The renal tissue specimens were evaluated for apoptosis and renal function using hematoxylin eosin staining and TUNEL staining. Changes in the emergence of active oxygen were investigated by using blood samples. The degree of renal dysfunction was evaluated by measuring neutrophil gelatinase-associated lipocalin (NGAL) in the spot urine samples. The changes in the serum creatinine level, showed that the renal function was preserved with a significant difference in the rhEPO administration group. The liver deviation enzymes clearly decreased in the rhEPO administration group. Active oxygen did not change before and after the ischemia-reperfusion nor was it changed by rhEPO administration. Apoptosis was inhibited by rhEPO administration. No direct effects of rhEPO administration on the emergence of active oxygen were observed. The administration of rhEPO, was suggested to help preserve the renal function in marginal donors with a longer agonal stageby effectively.

Topics: Animals; Apoptosis; Erythropoietin; In Situ Nick-End Labeling; Kidney; Kidney Transplantation; Male; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury

Acute systemic hypoxia induces delayed cardioprotection against ischaemia-reperfusion injury in the heart. As cobalt chloride (CoCl₂) is known to elicit hypoxia-like responses, it was hypothesized that this chemical would mimic the preconditioning effect and facilitate acclimatization to hypobaric hypoxia in rat heart.. Male Sprague-Dawley rats treated with distilled water or cobalt chloride (12.5 mg Co/kg for 7 days) were exposed to simulated altitude at 7622 m for different time periods (1, 2, 3 and 5 days).. Hypoxic preconditioning with cobalt appreciably attenuated hypobaric hypoxia-induced oxidative damage as observed by a decrease in free radical (reactive oxygen species) generation, oxidation of lipids and proteins. Interestingly, the observed effect was due to increased expression of the antioxidant proteins hemeoxygenase and metallothionein, as no significant change was observed in antioxidant enzyme activity. Hypoxic preconditioning with cobalt increased hypoxia-inducible factor 1α (HIF-1α) expression as well as HIF-1 DNA binding activity, which further resulted in increased expression of HIF-1 regulated genes such as erythropoietin, vascular endothelial growth factor and glucose transporter. A significant decrease was observed in lactate dehydrogenase activity and lactate levels in the heart of preconditioned animals compared with non-preconditioned animals exposed to hypoxia.. The results showed that hypoxic preconditioning with cobalt induces acclimatization by up-regulation of hemeoxygenase 1 and metallothionein 1 via HIF-1 stabilization.

Topics: Acclimatization; Animals; Cardiotonic Agents; Cobalt; DNA-Binding Proteins; Erythropoietin; Heart; Heme Oxygenase-1; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Male; Metallothionein; Myocardium; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Reperfusion Injury; Up-Regulation; Vascular Endothelial Growth Factor A

We investigated the protective effect of Erythropoietin (EPO) analogue rHuEPO on renal injury induced by acute exhaustive exercise in the rat. Rats were randomly allocated to one of 3 groups: normal control (C), exhaustive exercise test (ET) and EPO pre-treatment (rHuEPO 2 000 U/kg) plus ET (EPO+ET). Compared with controls, animals in the ET group had increased serum urea nitrogen, serum creatinine, urine protein, and renal tissue malondialdehyde (MDA) and decreased renal tissue nitric oxide (NO), nitric oxide synthase (NOS) and superoxide dismutase (SOD) activities. There was severe damage in renal tubular epithelial cells with a lot of cell apoptosis, and TUNEL assay revealed a remarkably high apoptotic index (p<0.01). Changes in renal function and kidney tissue were much less in the EPO+ET group (p<0.05) and the apoptotic index was much lower than in the ET group (18.45±0.32 vs. 27.55±0.49, p<0.05). EPO pretreatment thus significantly prevented renal cell apoptosis, and counteracted high MDA and low NO and NOS renal contents induced by exhaustive exercise. The data point to a potential value of EPO in preventing the acute renal injury after exhaustive exercise.

Topics: Acute Kidney Injury; Animals; Apoptosis; Erythropoietin; In Situ Nick-End Labeling; Male; Malondialdehyde; Nitric Oxide; Nitric Oxide Synthase; Physical Conditioning, Animal; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Ischemia-reperfusion injury (I/R) is the main cause of primary graft nonfunction. Our aim was to evaluate the effect of excessive versus acute administration of erythropoietin (EPO) in attenuating the hepatic injury induced by I/R in mice. The effect of segmental (70%) hepatic ischemia was evaluated in a transgenic mouse line with constitutive overexpression of human EPO cDNA and in wild-type (WT) mice. Mice were randomly allocated to 5 main experimental groups: (i) WT-sham, (ii) WT ischemia, (iii) WT ischemia + recombinant human erythropoietin (rhEPO), (iv) transgenic-sham, and (v) transgenic ischemia. The EPO-pretreated mice showed a significant reduction in liver enzyme levels and intrahepatic caspase-3 activity and fewer apoptotic hepatocytes (p < 0.05 for all) compared with the WT untreated I/R group. EPO decreased c-Jun N-terminal kinase (JNK) phosphorylation and nuclear factor-κB (NF-κB) expression during I/R. In transgenic I/R livers, baseline histology showed diffused hepatic injury, and no significant beneficial effect was noted between the WT untreated and the transgenic I/R mice. In conclusion, acute pretreatment with EPO in WT mice attenuated in vivo I/R liver injury. However, in excessive EPO overexpression, the initial liver injury abolished the beneficial effect of EPO. These findings have important implications for the potential use of acute EPO in I/R injury during liver transplantation.

Topics: Animals; Apoptosis; Caspase 3; DNA, Complementary; Erythropoietin; Female; Hematocrit; Hepatocytes; Humans; JNK Mitogen-Activated Protein Kinases; Liver; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; NF-kappa B; Phosphorylation; Random Allocation; Recombinant Proteins; Reperfusion Injury

Prothymosin-alpha (ProTalpha) causes a switch in cell death mode from necrosis to neurotrophin-reversible apoptosis in primary cultured cortical neurons. In the present study, post-ischemic administration (3 or 24 h, intravenously) of recombinant mouse ProTalpha without neurotrophins completely prevented ischemia-induced retinal damage accompanying necrosis and apoptosis, as well as dysfunction assessed by electroretinogram. Treatments with anti-erythropoietin (EPO) or brain-derived neurotrophic factor (BDNF) immunoglobulin G (IgG) reversed ProTalpha-induced inhibition of apoptosis. ProTalpha upregulated retinal EPO and BDNF levels in the presence of ischemia. Moreover, intravitreous administration of anti-ProTalpha IgG or an antisense oligodeoxynucleotide for ProTalpha accelerated ischemia-induced retinal damage. We also observed that ischemia treatment caused a depletion of ProTalpha from retinal cells. Altogether, these results suggest that the systemic administration of ProTalpha switches ischemia-induced necrosis to apoptosis, which in turn is inhibited by neurotrophic factors upregulated by ProTalpha and ischemia. ProTalpha released upon ischemic stress was found to have a defensive role in retinal ischemia.

Topics: Animals; Antibodies; Apoptosis; Brain-Derived Neurotrophic Factor; Erythropoietin; Ischemia; Male; Mice; Necrosis; Protein Precursors; Reperfusion Injury; Retina; Retinal Vessels; Thymosin

Erythropoietin (Epo) was recently defined as an endogenous agent with more than hematopoietic functions. Previously we explored the potential of this agent to ameliorate lung ischemia-reperfusion (I/R) injury. The present study aims to determine the optimal dose and timing of administration for improving lung injury, and to further investigate the mechanisms by which Epo ameliorates lung I/R injury. The left lungs of Sprague-Dawley rats underwent 90 min ischemia and 120 min reperfusion. Firstly, animals in different groups were intraperitoneally injected with various doses of recombined human erythropoietin (rhEpo) 24 h prior to operation, 2 h prior to operation, or after the onset of reperfusion. Pulmonary myeloperoxidase (MPO) activity and malondialdehyde (MDA) content were evaluated. Treatment with 3 KU/kg rhEpo 2 h prior to operation was optimal for attenuating pulmonary MPO activity and MDA content. With such treatment, ultrastructural changes of pneumocytes were observed, and the pneumocyte apoptosis index was also determined by terminal dUTP nick-end labeling method. The plasma concentrations of tumor necrosis factor (TNF)-alpha and matrix metalloproteinase (MMP)-9 were evaluated by enzyme-linked immunosorbent assay, and pulmonary expression by immunohistochemistry. When pretreated with rhEpo, the pneumocyte ultrastructure was predominantly maintained and the pulmonary apoptosis index was markedly reduced. In comparison with untreated animals, in treated animals the plasma concentrations of TNF-alpha and MMP-9 were significantly decreased, and their expression in lung tissue was markedly reduced as well. The results indicated that Epo potently protected against lung I/R injury by inhibiting systemic and local expression of TNF-alpha and MMP-9.

Topics: Animals; Apoptosis; Dose-Response Relationship, Drug; Erythropoietin; Gene Expression Regulation; Lung Injury; Male; Malondialdehyde; Matrix Metalloproteinase 9; Peroxidase; Pulmonary Alveoli; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Time Factors; Tumor Necrosis Factor-alpha

The mortality rate from acute kidney injury after major cardiovascular operations can be as high as 60%, and no therapies have been proved to prevent acute kidney injury in this setting. Here, we show that preconditioning with the anesthetic gas xenon activates hypoxia-inducible factor 1alpha (HIF-1alpha) and its downstream effectors erythropoietin and vascular endothelial growth factor in a time-dependent manner in the kidneys of adult mice. Xenon increased the efficiency of HIF-1alpha translation via modulation of the mammalian target of rapamycin pathway. In a model of renal ischemia-reperfusion injury, xenon provided morphologic and functional renoprotection; hydrodynamic injection of HIF-1alpha small interfering RNA demonstrated that this protection is HIF-1alpha dependent. These results suggest that xenon preconditioning is a natural inducer of HIF-1alpha and that administration of xenon before renal ischemia can prevent acute renal failure. If these data are confirmed in the clinical setting, then preconditioning with xenon may be beneficial before procedures that temporarily interrupt renal perfusion.

Topics: Anesthetics; Animals; Basic Helix-Loop-Helix Transcription Factors; Carrier Proteins; Cells, Cultured; Erythropoietin; Gene Expression Regulation; Humans; Ischemic Preconditioning; Kidney; Mice; Mice, Inbred C57BL; Phosphotransferases (Alcohol Group Acceptor); Protein Biosynthesis; Protein Subunits; Reperfusion Injury; RNA, Messenger; TOR Serine-Threonine Kinases; Transcription, Genetic; Ubiquitin; Vascular Endothelial Growth Factor A; Xenon

To observe the effect of recombinant human erythropoietin (rhEPO) on the expression of hypoxia inducible factor-1alpha (HIF-1alpha) in the retina of rabbits with acute high intraocular pressure and investigate the mechanism of rhEPO in protecting the retina from ischemia-reperfusion injury.. Acute high intraocular pressure was induced in the rabbits by perfusion of normal saline into the anterior chamber, and rhEPO was injected subcutaneously. The changes in HIF-1alpha protein expression in the retina was observed by immunohistochemistry on days 1, 3, 7, and 14 after retinal ischemia- reperfusion.. HIF-1alpha expression was not observed in the retina of the normal control rats, but intense HIF-1alpha expression was found in the model group (P<0.01). In rabbits with rhEPO injection and those in the model group, the patterns of HIF-1alpha expression alterations were similar, but the HIF-1alpha-positive cells in the retina were significantly fewer in rhEPO group (P<0.05).. rhEPO can down-regulate HIF-1alpha expression in the retina of rabbits with acute high intraocular pressure, which may be one of the mechanisms that rhEPO protects the retina from ischemia-reperfusion injury.

Topics: Animals; Down-Regulation; Erythropoietin; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Neuroprotective Agents; Ocular Hypertension; Rabbits; Recombinant Proteins; Reperfusion Injury; Retina; Retinal Vessels

Considering the implications that arose from several recent experimental studies using recombinant human erythropoietin in rodents, erythropoietin has been regarded as a pharmacological preconditioning agent. The purpose of the present study was to evaluate whether erythropoietin has a preconditioning effect against ischemia and reperfusion injury in the small intestine of the rat.. Intestinal ischemia was induced in male Wistar rats by clamping the superior mesenteric artery for 30 min, followed by reperfusion for 180 min. Recombinant human erythropoietin (1000 or 3000 U/kg) or vehicle was administered intraperitoneally 24 h prior to ischemia. After collection of ileal tissue, evaluation of damage was based on measurements of the accumulation of polymorphonuclear neutrophils by technetium-99m-labeled leukocyte uptake, content of malondialdehyde, reduced glutathione, contractile responses to agonists, and an evaluation of histopathological features in intestinal tissue.. Treatment with erythropoietin 24 h before ischemia significantly reduced the tissue content of malondialdehyde and increased that of reduced glutathione. Pretreatment also significantly suppressed leukocyte infiltration into the postischemic tissue, as evidenced by the lower content of myeloperoxidase and technetium-99m-labeled leukocytes. Physiological and histopathological improvements were also significant with the rHuEpo treatment.. Results of the present study indicate that rHuEpo is an effective preconditioning agent in ischemic injury of the small intestine. Protection provided by recombinant human erythropoietin is closely related to the inhibition of oxidative stress and leukocyte infiltration, which might be among the possible protective mechanisms of erythropoietin in intestinal ischemia and reperfusion.

Topics: Animals; Erythropoietin; Glutathione; Humans; Ileum; Injections, Intraperitoneal; Leukocytes; Male; Malondialdehyde; Muscle Contraction; Muscle, Smooth; Oxidative Stress; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury

Human recombinant Erythropoietin (rHuEpo) has recently been shown to be a potent protector of ischemia- reperfusion injury in warm-liver ischemia. Significant enhancement of hepatic regeneration and survival after large volume partial hepatic resection has also been demonstrated. It was the aim of this study to evaluate the capacities of rHuEpo in the setting of rat liver transplantation. One-hundred-and-twenty Wistar rats were used: 60 recipients received liver transplantation following donor organ treatment (60 donors) with either 1000 IU rHuEpo or saline injection (controls) into portal veins (cold ischemia 18 h, University of Wisconsin (UW) solution). Recipients were allocated to two groups, which either received 1000 IU rHuEpo at reperfusion or an equal amount of saline (control). Animals were sacrificed at defined time-points (2, 4.5, 24, 48 h and 7 days postoperatively) for analysis of liver enzymes, histology [hematoxylin-eosin (HE) staining, periodic acid Schiff staining (PAS)], immunostaining [terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), Hypoxyprobe] and real-time polymerase chain reaction (RT-PCR) of cytokine mRNA (IL-1, IL-6). Lactate dehydrogenase (LDH) and alanine aminotransferase (ALT) values were significantly reduced among the epo-treated animals 24 and 48 h after liver transplantation (LT). The TUNEL and Hypoxyprobe analyses as well as necrotic index evaluation displayed significant reduction of apoptosis and necrosis in rHuEpo-treated graft livers. Erythropoietin reduces ischemia-reperfusion injury after orthotopic liver transplantation in rats.

Topics: Animals; Apoptosis; Erythropoietin; Humans; Interleukin-1beta; Interleukin-6; Ischemia; Liver; Liver Transplantation; Rats; Rats, Wistar; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; Time Factors; Transplantation Conditioning

The purpose of this study was to examine the role of erythropoietin in retinal ischemic preconditioning (IPC).. Rats were subjected to retinal ischemia after IPC. Electroretinography assessed functional recovery after ischemia; retinal sections were examined to determine loss of retinal ganglion cells, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling was used to assess apoptosis. Levels of downstream mediators were measured in retinal homogenates by Western blotting. To assess the involvement of erythropoietin in IPC, Western blotting was used to measure levels of erythropoietin and its receptor (EPO-R) in retinal homogenates after IPC. To examine erythropoietin's role in IPC, the impact of blocking erythropoietin via intravitreal injection of soluble EPO-R (sEPO-R) before IPC was studied.. Erythropoietin levels did not change after IPC, but EPO-R increased. Intravitreal injection of sEPO-R significantly attenuated both the functional and histologic neuroprotection produced by IPC in comparison to control injection of denatured sEPO-R. Apoptotic damage after ischemia was enhanced in the sEPO-R-treated retinas as indicated by fluorescent terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling. Phosphorylated extracellular-signal-regulated kinase and heat shock protein 27, but not protein kinase B, upregulated in denatured sEPO-R-treated retinae, were attenuated in eyes injected with sEPO-R.. These results indicate that EPO-R upregulation is a critical component of the functional, histologic, and antiapoptotic protective effect of IPC on ischemia in the retina and that several downstream effectors may be involved in the neuroprotective actions of erythropoietin.

Topics: Animals; Apoptosis; Blotting, Western; DNA Nucleotidylexotransferase; Electroretinography; Erythropoietin; Extracellular Signal-Regulated MAP Kinases; Fluorescent Antibody Technique, Indirect; HSP27 Heat-Shock Proteins; In Situ Nick-End Labeling; Ischemic Preconditioning; Neurons; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Receptors, Erythropoietin; Reperfusion Injury; Retina; Retinal Vessels; Solubility

To evaluate the effects of erythropoietin (EPO) as an antioxidant and tissue protective agent and study the biochemical and histopathological changes in experimental ischemia and ischemia/reperfusion (I/R) injury in rat ovaries.. 36 Adult female rats were used. The experimental groups were designed as Group 1: sham operation; Group 2: bilateral ovarian ischemia; and Group 3: 3 h period of ischemia followed by 3 h reperfusion. Group 4 rats were administered a 5000 IU dose of EPO, before 0.5 h of ischemia, and then bilateral ovarian ischemia was applied. After a 3 h period of ischemia, the bilateral ovaries were removed. In Group 5, a 3 h period of bilateral ovarian ischemia was applied. 2.5 h after the induction of ischemia, the rats were administered the same dose of EPO. At the end of a 3 h period of ischemia, 3h reperfusion was continued after the ovaries were removed. Group 6 underwent a sham operation after administration of 5000 IU/kg of EPO. After the experiments, superoxide dismutase (SOD), inducible nitric oxide synthase (iNOS), and myeloperoxidase (MPO) activity were determined, and histopathological changes were examined in all rat ovarian tissue.. Ischemia and ischemia/reperfusion increased the iNOS and MPO activity while decreasing the SOD activity significantly in comparison to the sham group. The 5000 IU/kg of EPO before ischemia and I/R reversed the trend in iNOS and MPO activities. The levels of SOD were decreased by the ischemia and I/R. The administration of EPO before ischemia and I/R treatments also reversed the trend in the SOD levels. In the ischemia/reperfusion plus EPO groups, though we observed minimal vascular dilation in the ovary stroma and some degenerative cell clusters, most of cellular structures did not show any pathological changes.. Administration of EPO is effective in reversing tissue damage induced by ischemia and/or ischemia/reperfusion in ovaries.

Topics: Animals; Antioxidants; Erythropoietin; Female; Ischemia; Ovarian Diseases; Ovary; Rats; Rats, Wistar; Reperfusion Injury

Recombinant human erythropoietin alpha (rHu-EPO) has been reported to protect the liver of rats and mice from ischemia-reperfusion injury. However, direct protective effects of rHu-EPO on hepatocytes and the responsible signalling pathways have not yet been described. The aim of the present work was to study the protective effect of rHu-EPO on warm hypoxia-reoxygenation and cold-induced injury to hepatocytes and the rHu-EPO-dependent signalling involved.. Loss of viability of isolated rat hepatocytes subjected to hypoxia/reoxygenation or incubated at 4 degrees C followed by rewarming was determined from released lactate dehydrogenase activity in the absence and presence of rHu-EPO (0.2-100 U/ml). Apoptotic nuclear morphology was assessed by fluorescence microscopy using the nuclear fluorophores H33342 and propidium iodide. Erythropoietin receptor (EPOR), EPO and Bcl-2 mRNAs were quantified by real time PCR. Activation of JAK-2, STAT-3 and STAT-5 in hepatocytes and rat livers perfused in situ was assessed by Western blotting.. In contrast to previous in vivo studies on ischemia-reperfusion injury to the liver, rHu-EPO was without any protective effect on hypoxic injury, hypoxia-reoxygenation injury and cold-induced apoptosis to isolated cultured rat hepatocytes. EPOR mRNA was identified in these cells but specific detection of the EPO receptor protein was not possible due to the lack of antibody specificity. Both, in the cultured rat hepatocytes (10 U/ml for 15 minutes) and in the rat liver perfused in situ with rHu-EPO (8.9 U/ml for 15 minutes) no evidence for EPO-dependent signalling was found as indicated by missing effects of rHu-EPO on phosphorylation of JAK-2, STAT-3 and STAT-5 and on the induction of Bcl-2 mRNA.. Together, these results indicate the absence of any protective EPO signalling in rat hepatocytes. This implies that the protection provided by rHu-EPO in vivo against ischemia-reperfusion and other causes of liver injury is most likely indirect and does not result from a direct effect on hepatocytes.

Topics: Animals; Apoptosis; Cell Hypoxia; Cell Line; Cells, Cultured; Cold Temperature; Erythropoietin; Hematopoietic Stem Cells; Hepatocytes; Janus Kinase 2; Male; Models, Animal; Proto-Oncogene Proteins c-bcl-2; Rats; Recombinant Proteins; Reperfusion Injury; RNA, Messenger; Signal Transduction; STAT3 Transcription Factor; STAT5 Transcription Factor

Ischemia/reperfusion (I/R) injury is a major cause of acute renal failure in kidney transplantation; however, the mechanisms of kidney damage and repair are not yet clear. So far no treatment has been effective to prevent I/R injury. In the present study we evaluated the effect of erythropoetin (EPO) in I/R injury in rats. We investigated the role of bone marrow cells (BMC) in kidney repair and the effect of EPO on BMC recruitment.. Female Sprague Dawley rats transplanted with male BMCs underwent I/R injury. In the treatment group rats received 5000 IU of EPO 30 minutes before renal ischemia. At 2 and 4 weeks after I/R, rats were humanely killed and we measured creatinine clearance (glomerular filtration rate [GFR]), proteinuria, and body weight (BW). Renal tissue was harvested for histologic and molecular analysis. Fluorescein in situ hybridization (FISH) and TUNEL were used to determined the presence of male cell chimerism and apoptosis in renal tissue.. At 4 weeks after I/R, EPO significantly improved GFR (1.8 +/- 0.2 vs 1.2 +/- 0.14 mL/min; P < .05). No significant differences between EPO and control rats were observed in proteinuria, BW, and hemoglobin levels at 2 and 4 weeks. After death, the kidney showed only minimal tubulointerstitial changes, which were more marked in control rats. FISH analysis demonstrated a low degree of microchimerism, not significantly different between EPO and control rats. Apoptosis decreased between 2 and 4 weeks after I/R, in both EPO and control groups.. EPO improved GFR and injury at 4 weeks after I/R; however, it did not enhance the recruitment of BMC.

Topics: Animals; Apoptosis; Bone Marrow Cells; Bone Marrow Transplantation; Erythropoietin; Female; Glomerular Filtration Rate; In Situ Hybridization, Fluorescence; In Situ Nick-End Labeling; Kidney; Male; Rats, Sprague-Dawley; Reperfusion Injury

During hepatectomy, a period of ischemia and restoration of the blood supply can result in hepatic ischemia-reperfusion injury (IRI). Current research indicates that erythropoietin (EPO) has a protective effect in animal models of cerebral ischemia, myocardial infarction, and renal IRI. However there is lack of research into the role of EPO in hepatic IRI. This study aimed to explore the role of EPO in hepatic IRI and its possible mechanism of action.. Thirty male Sprague-Dawley rats were divided into three groups: (1) ten rats in the experimental group were given 1000 IU/kg EPO one day before the operation; (2) ten rats in a control group were given normal saline preoperatively as a placebo; and (3) ten rats served as a sham-operated group. Hepatic IRI was induced by occluding the hepatic arteries of the three cephalad hepatic segments and the portal vein for about 45 minutes, while in the sham-operated group only laparotomy was performed. The levels of ALT and AST were tested 24 hours pre- and post-operation. All rats were sacrificed 24 hours after the operation to assess the pathologic changes in the liver and measure the expression of heme oxygenase-1 (HO-1) through Western blotting and RT-PCR.. Hepatic IRI was markedly mitigated in the experimental group as compared with the control group. Moreover, the expression of HO-1 at the level of both transcription and protein increased prominently (P<0.05) in the experimental group.. These results demonstrate that EPO can up-regulate HO-1 in liver tissues and accordingly decrease hepatic injury through its anti-inflammatory property.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Blotting, Western; Erythropoietin; Heme Oxygenase-1; Hepatectomy; Liver; Male; Postoperative Period; Preoperative Care; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Up-Regulation

Renal ischemia and reperfusion injury leads to acute renal failure when proinflammatory and apoptotic processes in the kidney are activated. The increase in hypoxia-inducible transcription factor-alpha (HIF-alpha), an important transcription factor for several genes, can attenuate ischemic renal injury. We recently identified a novel WD-repeat protein designated Morg1 (MAPK organizer 1) that interacts with prolyl hydroxylase 3 (PHD3), an important enzyme involved in the regulation of HIF-1alpha and HIF-2alpha expression. While homozygous Morg1 -/- mice are embryonic lethal, heterozygous Morg1 +/- mice have a normal phenotype. We show here that Morg1 +/- were partially protected from renal ischemia-reperfusion injury compared with wild-type Morg1 +/+ animals. Morg1 +/- mice compared with wild-type animals revealed a stronger increase in HIF-1alpha and HIF-2alpha expression in the ischemic-reperfused kidney associated with enhanced serum erythropoietin levels. However, no significant expression of HIF-1alpha and HIF-2alpha was found in nonischemic kidneys without any difference between Morg1 +/- and Morg1 +/+ mice. Ischemic kidneys of Morg1 +/- mice expressed more erythropoietin mRNA than ischemic kidneys from wild-type animals. Renal ischemia in Morg1 +/- mice resulted in a decrease in renal inflammation and reduction of proinflammatory cytokines (MCP-1, IP-10, MIP-2) compared with wild-type mice. Furthermore, there was significantly less apoptosis and tubular damage in Morg1 +/- kidneys after ischemia-reperfusion, and this was also reflected in significantly improved renal function compared with wild-type. Thus Morg1 may be a novel therapeutic target to limit renal injury after ischemia-reperfusion.

Topics: Acute Disease; Adaptor Proteins, Signal Transducing; Animals; Apoptosis; Blotting, Western; DNA, Complementary; Electrophoretic Mobility Shift Assay; Erythropoietin; Female; Heterozygote; Hypoxia-Inducible Factor 1, alpha Subunit; Immunohistochemistry; Kidney; Kidney Diseases; Kidney Function Tests; Mice; Mice, Inbred C57BL; Mice, Knockout; Pregnancy; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; RNA

The aim of the study is to investigate the effects of erythropoietin on torsion/detorsion injury in rats.. Forty rats were divided randomly into 5 groups: group I (sham, S), sham operation; group II (torsion/detorsion 1, T/D(1)), 3 hours ischemia and 1 hour reperfusion; group III (torsion/detorsion 2, T/D(2)), 3 hours ischemia and 48 hours reperfusion; group IV (erythropoietin 1, EPO(1)), 3 hours ischemia, 1 hour reperfusion, and a single dose of EPO; and group V (erythropoietin 2, EPO(2)), 3 hours ischemia, 48 hours reperfusion, and 2 doses of EPO. Malondialdehyde (MDA) and nitric oxide (NO) levels and activities of superoxide dismutase and catalase were measured. Tissue damage to ovarian tissue was scored by histologic examination. Data were compared among groups with parametric tests.. The MDA levels in the S and EPO groups were significantly lower than the T/D groups (P < .001). Catalase and superoxide dismutase activities, and NO levels in the S and EPO groups were significantly higher than in the T/D groups (P < .05). Ovarian tissue damage in the S and EPO groups was significantly less than in the T/D groups (P < .05). Levels of all biochemical markers and ovarian tissue damage scores were similar among the S, EPO(1), and EPO(2) groups (P > .05).. Erythropoietin attenuates ischemia-reperfusion injury when given during the acute phase of ovarian torsion-detorsion in a rat model.

Topics: Animals; Antioxidants; Catalase; Disease Models, Animal; Erythropoietin; Female; Malondialdehyde; Nitric Oxide; Ovarian Diseases; Ovary; Random Allocation; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Superoxide Dismutase; Torsion Abnormality

To investigate the protective effects of ginkgolide B and hypoxic preconditioning against acute hypoxia injury in mice.. Ordinary pressure acute hypoxia model in mice was adopted to observe the ethology, the duration of the death and the degree of brain edema. Meanwhile the expression of RTP801 mRNA and erythropoietin (EPO) were measured by RT-PCR and Western blot, respectively.. Ginkgolide B and hypoxic preconditioning could both prolong the survival time of hypoxia under ordinary pressure,and significantly decreased the degree of brain edema. Besides ginkgolide B and hypoxic preconditioning could both up-regulate the expression of RTP801mRNA and EPO.. Ginkgolide B has the similar effect to hypoxic preconditioning against acute hypoxia. Both of these protective effects may be associated with the up-regulation of the expression of RTP801 mRNA and EPO.

Topics: Animals; Brain; Brain Edema; Erythropoietin; Female; Ginkgolides; Hypoxia; Ischemic Preconditioning; Lactones; Male; Mice; Mice, Inbred ICR; Reperfusion Injury; Repressor Proteins; RNA, Messenger; Transcription Factors; Up-Regulation

Aortic occlusion causes ischemia/reperfusion injury, kidney and spinal cord being the most vulnerable organs. Erythropoietin improved ischemia/reperfusion injury in rodents, which, however, better tolerate ischemia/reperfusion than larger species. Therefore, we investigated whether erythropoietin attenuates porcine aortic occlusion ischemia/reperfusion injury.. Before occluding the aorta for 45 mins by inflating intravascular balloons, we randomly infused either erythropoietin (n = 8; 300 IU/kg each over 30 mins before and during the first 4 hrs of reperfusion) or vehicle (n = 6). During aortic occlusion, mean arterial pressure was maintained at 80% to 120% of baseline by esmolol, nitroglycerine, and adenosine 5'-triphosphate. During reperfusion, noradrenaline was titrated to keep mean arterial pressure >80% of baseline. Kidney perfusion and function were assessed by fractional Na-excretion, p-aminohippuric acid and creatinine clearance, spinal cord function by lower extremity reflexes and motor evoked potentials. Blood isoprostane levels as well as blood and tissue catalase and superoxide dismutase activities allowed evaluation of oxidative stress. After 8 hrs of reperfusion, kidney and spinal cord specimens were taken for histology (hematoxylin-eosin, Nissl staining) and immunohistochemistry (TUNEL assay for apoptosis).. Parameters of oxidative stress and antioxidative activity were comparable. Erythropoietin reduced the noradrenaline requirements to achieve the hemodynamic targets and may improve kidney function despite similar organ blood flow, histology, and TUNEL staining. Neuronal damage and apoptosis was attenuated in the thoracic spinal cord segments without improvement of its function.. During porcine aortic occlusion-induced ischemia/reperfusion erythropoietin improved kidney function and spinal cord integrity. The lacking effect on spinal cord function was most likely the result of the pronounced neuronal damage associated with the longlasting ischemia.

Topics: Animals; Arterial Occlusive Diseases; Balloon Occlusion; Disease Models, Animal; Erythropoietin; Evoked Potentials, Motor; Female; Hemodynamics; Kidney Function Tests; Male; Oxidative Stress; Reperfusion Injury; Spinal Cord Injuries; Swine

Topics: Animals; Down-Regulation; Erythropoietin; Gene Expression; Intercellular Adhesion Molecule-1; Lymphocyte Function-Associated Antigen-1; Neutrophil Activation; Rats; Recombinant Proteins; Reperfusion Injury; Surgical Flaps

Renal injury induced by aortic ischemia-reperfusion (IR) is an important factor in the development of postoperative acute renal failure following abdominal aortic surgery. The purpose of this study is to examine the effect of erythropoietin on renal injury induced by aortic IR in rats.. Twenty-four Wistar-Albino rats were randomized into 3 groups (8 per group). The control group underwent laparotomy and dissection of the infrarenal abdominal aorta without occlusion. The aortic IR group underwent clamping of the infrarenal abdominal aorta for 30 min followed by 60 min of reperfusion. The aortic IR + erythropoietin group underwent the same aortic IR periods and was pretreated with 1000 U/kg subcutaneous erythropoietin 5 min before ischemia. In rat kidney specimens, tissue levels of malondialdehyde (MDA), superoxide dismutase, catalase, and glutathione peroxidase were measured. Histological evaluation of the rat kidney tissues was also done.. Aortic IR significantly increased the levels of MDA and superoxide dismutase (P < 0.05 versus control). Erythropoietin significantly decreased the levels of MDA, superoxide dismutase, and catalase (P < 0.05 versus aortic IR). Histological evaluation showed that aortic IR significantly increased (P < 0.05 versus control), whereas erythropoietin significantly decreased (P < 0.05 versus aortic IR) the focal glomerular necrosis, dilation of Bowman's capsule, degeneration of tubular epithelium, necrosis in tubular epithelium, interstitial inflammatory infiltration, and congestion of blood vessels.. The results indicate that erythropoietin has protective effects on renal injury induced by aortic IR in rats.

Topics: Acute Kidney Injury; Animals; Aorta, Abdominal; Catalase; Erythropoietin; Female; Glutathione Peroxidase; Kidney; Male; Malondialdehyde; Rats; Rats, Wistar; Reperfusion Injury; Superoxide Dismutase

Topics: Animals; Disease Models, Animal; Erythropoietin; Humans; Neuroprotective Agents; Reperfusion Injury; Species Specificity; Spinal Cord Ischemia

Erythropoietin (EPO), a member of the type 1 cytokine superfamily, plays a critical hormonal role regulating erythrocyte production as well as a paracrine/autocrine role in which locally produced EPO protects a wide variety of tissues from diverse injuries. Significantly, these functions are mediated by distinct receptors: hematopoiesis via the EPO receptor homodimer and tissue protection via a heterocomplex composed of the EPO receptor and CD131, the beta common receptor. In the present work, we have delimited tissue-protective domains within EPO to short peptide sequences. We demonstrate that helix B (amino acid residues 58-82) of EPO, which faces the aqueous medium when EPO is bound to the receptor homodimer, is both neuroprotective in vitro and tissue protective in vivo in a variety of models, including ischemic stroke, diabetes-induced retinal edema, and peripheral nerve trauma. Remarkably, an 11-aa peptide composed of adjacent amino acids forming the aqueous face of helix B is also tissue protective, as confirmed by its therapeutic benefit in models of ischemic stroke and renal ischemia-reperfusion. Further, this peptide simulating the aqueous surface of helix B also exhibits EPO's trophic effects by accelerating wound healing and augmenting cognitive function in rodents. As anticipated, neither helix B nor the 11-aa peptide is erythropoietic in vitro or in vivo. Thus, the tissue-protective activities of EPO are mimicked by small, nonerythropoietic peptides that simulate a portion of EPO's three-dimensional structure.

Topics: Animals; Cytokine Receptor Common beta Subunit; Erythropoietin; Kidney; Male; Mice; Mice, Inbred C57BL; Papilledema; Pattern Recognition, Visual; Peptides; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Wound Healing

Administration of erythropoietin (EPO) during or immediately after myocardial ischemia can reduce subsequent myocardial apoptosis, a key phenomenon in myocardial ischemia-reperfusion injury. In this study, we assessed the effect of EPO on (99m)Tc-annexin V myocardial uptake and whether the accumulation of (99m)Tc-annexin V can predict cardiac remodeling and functional deterioration.. Eighteen rats with left coronary artery (LCA) occlusion were randomized to receive either an intravenous injection of EPO (EPO group) or saline (nontherapy [nT] group) immediately after release of the occlusion. After 20 min of LCA occlusion and 30 min of reperfusion, the rats were injected with (99m)Tc-annexin V. One hour after (99m)Tc-annexin V injection, the LCA was reoccluded and (201)Tl was injected intravenously, and the rats were sacrificed 1 min later. The heart was removed and sectioned, and dual-tracer autoradiography was performed to evaluate the distribution of the area at risk (defined on the thallium autoradiograph) and the area of apoptosis (defined on the annexin autoradiograph). Adjacent histologic specimens had deoxyuridine triphosphate nick-end labeling (TUNEL) staining to confirm the presence of apoptosis and were compared with autoradiography. Another 16 rats were randomized to EPO and nT groups and underwent echocardiography immediately after release of the LCA occlusion and at 2 and 4 wk after surgery.. The areas of (99m)Tc-annexin V accumulation in the EPO group were smaller than those in the nT group, though the (201)Tl defect areas of these 2 groups were comparable (area ratio, 0.318 +/- 0.038 vs. 0.843 +/- 0.051, P < 0.001, for annexin and 24.8 +/- 2.1 vs. 25.9 +/- 2.6 mm(2), P = NS, for thallium). (99m)Tc-annexin V accumulation correlated with the density of TUNEL-positive cells (r = 0.886, P < 0.001). In the nT group, left ventricular end-diastolic dimension (Dd) increased from baseline at 2 wk by 34.7% +/- 3.8% and remained stable at 34.9% +/- 5.0% at 4 wk after coronary occlusion. In the EPO group, Dd increased by 8.5% +/- 2.1% (P < 0.01 vs. nT at 2 wk) and 13.2% +/- 2.8% (P < 0.01 vs. nT at 4 wk). In the nT group, the left ventricular percentage of fractional shortening decreased by 42.2% +/- 3.4% and 52.9% +/- 3.4% at 2 and 4 wk, respectively, whereas in the EPO group it decreased 9.0% +/- 1.9% at 2 wk (P < 0.01 vs. nT at 2 wk) and 11.1% +/- 6.7% at 4 wk (P < 0.01 vs. nT at 4 wk).. This study demonstrated that a single treatment with EPO immediately after release of coronary ligation suppressed cardiac remodeling and functional deterioration. (99m)Tc-annexin V autoradiographs and TUNEL staining confirm that this change is due to a decrease in the extent of myocardial apoptosis in the ischemic/reperfused region.

Topics: Animals; Annexin A5; Apoptosis; Autoradiography; Cardiotonic Agents; Coronary Vessels; Echocardiography; Erythropoietin; Hematocrit; Male; Myocardial Infarction; Radionuclide Imaging; Rats; Rats, Wistar; Reperfusion Injury; Technetium

Topics: Erythropoietin; Humans; Reperfusion Injury

Preconditioning, such as by brief hypoxic exposure, has been shown to protect hearts against severe ischaemia. Here we hypothesized that hypoxic preconditioning (HPC) protects injured hearts by mobilizing the circulating progenitor cells. Ischaemia-reperfusion (IR) injury was induced by left coronary ligation and release in rats kept in room air or preconditioned with 10% oxygen for 6 weeks. To study the role of erythropoietin (EPO), another HPC + IR group was given an EPO receptor (EPOR) antibody via a subcutaneous mini-osmotic pump 3 weeks before IR induction. HPC alone gradually increased haematocrit, cardiac and plasma EPO, and cardiac vascular endothelial growth factor (VEGF) only in the first two weeks. HPC improved heart contractility, reduced ischaemic injury, and maintained EPO and EPOR levels in the infarct tissues of IR hearts, but had no significant effect on VEGF. Interestingly, the number of CD34(+)CXCR4(+) cells in the peripheral blood and their expression in HPC-treated hearts was higher than in control. Preconditioning up-regulated cardiac expression of stromal derived factor-1 (SDF-1) and prevented its IR-induced reduction. The EPOR antibody abolished HPC-mediated functional recovery, and reduced SDF-1, CXCR4 and CD34 expression in IR hearts, as well as the number of CD34(+)CXCR4(+) cells in blood. The specificity of neutralizing antibody was confirmed in an H9c2 culture system. In conclusion, exposure of rats to moderate hypoxia leads to an increase in progenitor cells in the heart and circulation. This effect is dependent on EPO, which induces cell homing by increased SDF-1/CXCR4 and reduces the heart susceptibly to IR injury.

Topics: Animals; Antibodies; Antigens, CD34; Chemokine CXCL12; Creatine Kinase, MB Form; Erythropoietin; Gene Expression; Heart; Hypoxia; Ischemic Preconditioning, Myocardial; Male; Models, Biological; Myocardium; Myocytes, Cardiac; Rats; Rats, Wistar; Receptors, CXCR4; Receptors, Erythropoietin; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; Stem Cells; Vascular Endothelial Growth Factor A; Ventricular Function, Left

To investigate the anti-inflammatory effect of erythropoietin (EPO) pretreatment on cardiomyocytes exposed to hypoxia/reoxygenation injury (H/R) and explore the possible mechanism.. The cultured neonatal rats?ventricular cardiomyocytes were divided randomly into 4 groups, control group (C group), EPO pretreatment group (E group), EPO and pyrrolidine dithiocarbamate (PDTC) pretreatment group (EP group) and PDTC pretreatment group (P group). After 24 hours?pretreatment, the cardiomyocytes were exposed to H/R. After pretreatment and H/R, the expression of tumor necrosis factor-alpha(TNF-alpha) gene in all the groups was detected by RT-PCR and Western blot. The nuclear factor-kappa B (NF-kappa B) activity was detected by electrophoretic mobility shift assay (EMSA) and the inhibitor-kappa B alpha (I-kappa B alpha) protein level was detected by Western blot.. The decrement of I-kappa B alpha protein and the increasing NF-kappa B activity were found in cardiomyocytes pretreated with EPO before H/R compared to other groups (t equal to 3.321, 4.183, P less than 0.01). However, after H/R, NF-kappa B activity and expression of TNF-alphagene were significantly reduced, I-kappa B alpha protein expression was increased in cardiomyocytes of E group compared to other groups (t=3.425, 3.687, 3.454, P less than 0.01). All theses changes caused by EPO pretreatment were eliminated by the intervention of PDTC (an antagonist to NF-kappa B) during pretreatment.. EPO pretreatment can inhibit the activation of NF-kappa B and upregulation of TNF-alpha gene in cardiomyocytes exposed to H/R through a negative feedback of NF-kappa B signaling pathway, and thus produces the anti-inflammatory effect. This might be one of the ways EPO produces the anti-inflammatory effect.

Topics: Analysis of Variance; Animals; Animals, Newborn; Antioxidants; Blotting, Western; Cells, Cultured; Electrophoretic Mobility Shift Assay; Erythropoietin; Hypoxia; Inflammation; Myocytes, Cardiac; NF-kappa B; Pyrrolidines; Random Allocation; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; Thiocarbamates; Tumor Necrosis Factor-alpha

To investigate the effects of erythropoietin (EPO) on the expression of aquaporin 2 (AQP(2)) after renal ischemia/reperfusion (IR).. Twenty-four Wistar rats were randomly divided into 3 equal groups: IR group undergoing resection of the right kidney, closuring of the left renal artery, vein, and ureter, and un-closuring 40 min later; IR + EPO group undergoing the above mentioned procedures and then intraperitoneal injection of EPO 3000 U/kg on days 1 and 2 after the treatment; and control group undergoing resection of the right kidney only without IR of the left kidney. Urine volume and urine osmotic pressure were measured. Blood samples were collected to detect the serum blood urea nitrogen (BUN) and creatinine (Cr). Three days after the treatment the kidneys were taken out. RT-PCR and Western blotting were used to detect the mRNA and protein expression of AQP(2).. The urine volume of the IR + EPO group was (26.0 +/- 2.3) microl .min(-1).kg(-1), significantly lower than that of the IR group [(59.1 +/- 1.3) microl .min(-1) . kg(-1), P < 0.01]. The urine osmotic pressure of the IR + EPO group was (1508 +/- 121) mOsm/kg H(2)O, significantly higher than that of the IR group [(235 +/- 99) mOsm/kg H(2)O, P < 0.01]. The serum BUN and Cr levels of the IR + EPO group were (12.3 +/- 6.0) mmol/L and (51 +/- 5) micromol/L respectively, both significantly lower than those of the IR group [(29.9 +/- 3.7) mmol/L and (141 +/- 5) micromol/L respectively, both P < 0.01]. The mRNA and protein expression of AQP(2) were highly positive in the control group. The protein expression levels of AQP(2) of the IR + EPO group were not significantly different from those of the control group (both P > 0.05), and the protein expression levels of AQP(2) of the IR group were significantly lower than those of the control group (both P < 0.01).. EPO can inhibit the down-regulation of AQP(2) in response to IR and this may take part in the EPO protective mechanism of renal ischemia/reperfusion injury.

Topics: Animals; Aquaporin 2; Disease Models, Animal; Erythropoietin; Female; Kidney; Kidney Diseases; Rats; Rats, Wistar; Reperfusion Injury

Recent studies have shown that insulin growth factor-1 (IGF-1) and either erythropoietin (EPO) or the long-acting EPO analog Darbepoetin alfa (DA) protect the heart against ischemia/reperfusion (I/R) and myocardial infarction (MI). The present study examined the cardioprotective effect of simultaneous treatments with IGF-1 and DA in these models of cardiac injury. Rats were subjected to I/R or MI and were treated with IGF-1, DA, and a combination of IGF-1 and DA, or vehicle treatment. IGF-1 and DA treatments imparted similar protective effect by reducing infarct size. Moreover, these treatments led to improvement of cardiac function after I/R or MI compared to vehicle. In the reperfused heart, apoptosis was reduced with either or both IGF-1 and DA treatments as measured by reduced TUNEL staining and caspase-3 activity. In addition, after MI, treatment with IGF-1 or DA significantly induced angiogenesis. This angiogenic effect was enhanced significantly when IGF-1 and DA were given simultaneously compared to vehicle or either agents alone. These data indicate simultaneous pharmacological treatments with IGF-1 and DA protect the heart against I/R and MI injuries. This protection results in reduced infarct size and improved cardiac function. Moreover, this treatment reduces apoptosis and enhances angiogenesis in the ischemic heart.

Topics: Animals; Apoptosis; Caspase 3; Darbepoetin alfa; Erythropoietin; In Situ Nick-End Labeling; Insulin-Like Growth Factor I; Male; Models, Biological; Myocardial Infarction; Myocardial Ischemia; Myocardium; Neovascularization, Pathologic; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Recombinant human erythropoietin (rhEpo) has been shown to reduce tissue injury following ischemia-reperfusion. We examined whether rhEpo protects in vitro renal tubular epithelial cells against radiocontrast media-induced injury.. LLC-PK1 renal tubular epithelial cells were exposed to non-ionic radiocontrast agent iohexol (low-osmolar) or iodixanol (iso-osmolar), with or without rhEpo (200 U/ml). Following a 6-hour exposure, cells were incubated for 24 h in radiocontrast-free culture medium. Cell viability was then assessed by the MTT assay. We also assessed cell apoptosis by the TUNEL assay, and activities of caspase-3, caspase-8, and caspase-9 were determined by a luminescence assay.. rhEpo improved viability of iohexol-treated LLC-PK1 cells by 27 +/- 6% (88.1 +/- 1.5 vs. 70.8 +/- 3.3%, p = 0.008). Similarly, rhEpo improved the viability of iodixanol-treated LLC-PK1 cells by 26 +/- 4% (82.5 +/- 2.1vs. 65.7 +/- 1.7%, p = 0.028). rhEpo also decreased apoptosis rates of iohexol-treated LLC-PK1 cells (6.4 +/- 0.9/1,000 cells vs. 14.8 +/- 2.4/1,000 cells, p = 0.028), and iodixanol-treated LLC-PK1 cells (8.0 +/- 1.2/1,000 cells vs. 13.5 +/- 1.9/1,000 cells, p = 0.028). In iohexol-treated LLC-PK1 cells, rhEpo attenuated activation of caspase-3 (p = 0.003), caspase-8 (p = 0.033) and caspase-9 (p = 0.055).. rhEpo attenuates in vitro renal tubular epithelial cell injury induced by low- and iso-osmolar radiocontrast media, possibly by reduction of caspases activation and apoptosis rates.

Topics: Animals; Apoptosis; Caspases; Cell Survival; Contrast Media; Enzyme Activation; Epithelial Cells; Erythropoietin; In Situ Nick-End Labeling; Iohexol; Kidney Tubules; LLC-PK1 Cells; Recombinant Proteins; Reperfusion Injury; Swine; Triiodobenzoic Acids

Apoptosis is a central mechanism of cell death following reperfusion of the ischemic liver. Recombinant human erythropoietin (rhEPO) have an important role in the treatment of myocardial ischemia/reperfusion (I/R) injury, by preventing apoptosis. The aim of the study was to investigate the effect of different regimens of rhEPO in preventing apoptosis following I/R-induced hepatic injury.. Isolated mouse livers were randomly divided into five groups: (1) control group, perfused for the whole study period (105 min); (2) 30-min perfusion followed by 90 min of ischemia and 15 min of reperfusion; (3), (4) and (5) like group 2, but with administration of rhEPO 5,000 units/kg i.p. at 30 min, 24 h, or both 30 min and 24 h respectively, before induction of ischemia. Perfusate liver enzyme levels and intrahepatic caspase-3 activity were measured, and apoptotic cells were identified by morphological criteria, TUNEL assay, and immunohistochemistry for caspase-3. Using immunoblot the expression of the proapoptotic JNK and inhibitor of NFkappaB (IkappaBalpha) were also evaluated. von Willebrand factor (vWF) immunohistochemistry was used as a marker of endothelial cells.. Compared to the I/R livers, all 3 rhEPO pretreated groups showed: a significant reduction in liver enzyme levels (P < 0.05) and intrahepatic caspase-3 activity (P < 0.05), fewer apoptotic hepatocytes (P < 0.05) and positive vWF staining in numerous endothelial cells lining the sinusoids. EPO decreased JNK phosphorylation and the degradation of the inhibitor of NFkappaB (IkappaBalpha) during I/R. There was no added benefit of the multiple- over the single-dose rhEPO regimen.. Pretreatment with one dose of rhEPO can attenuate post-I/R hepatocyte apoptotic liver damage. NFkappaB and JNK activation is likely to play a pivotal role in the pathophysiology of I/R hepatic injury and might have a key role in EPO-mediated protective effects. This effect is associated with the increase in sinusoidal vWF immunostaining suggests an additional effect of rhEPO in liver angiogenesis recovery. These findings have important implications for the potential use of rhEPO in I/R injury during liver transplantation.

Topics: Animals; Apoptosis; Caspase 3; Disease Models, Animal; Erythropoietin; Humans; Ischemia; Liver; Male; MAP Kinase Kinase 4; Mice; NF-kappaB-Inducing Kinase; Protein Serine-Threonine Kinases; Random Allocation; Recombinant Proteins; Reperfusion; Reperfusion Injury; von Willebrand Factor

We studied the effect of hyperbaric oxygen (HBO) preconditioning on the molecular mechanisms of neuroprotection in a rat focal cerebral ischemic model. Seventy-two male Sprague-Dawley rats were pretreated with HBO (100% O(2), 2 atmospheres absolute, 1 h once every other day for 5 sessions) or with room air. In experiment 1, HBO-preconditioned rats and matched room air controls were subjected to focal cerebral ischemia or sham surgery. Postinjury motor parameters and infarction volumes of HBO-preconditioned rats were compared with those of controls. In experiment 2, HBO-preconditioned rats and matched room air controls were killed at different time points. Brain levels of hypoxia-inducible factor-1alpha (HIF-1alpha) and its downstream target gene erythropoietin (EPO) analyzed by Western blotting and RT-PCR as well as HIF-1alpha DNA-binding and transcriptional activities were determined in the ipsilateral hemisphere. HBO induced a marked increase in the protein expressions of HIF-1alpha and EPO and the activity of HIF-1alpha, as well as the expression of EPO mRNA. HBO preconditioning dramatically improved the neurobehavioral outcome at all time points (3.0 +/- 2.1 vs. 5.6 +/- 1.5 at 4 h, 5.0 +/- 1.8 vs. 8.8 +/- 1.4 at 8 h, 6.4 +/- 1.8 vs. 9.7 +/- 1.3 at 24 h; P < 0.01, respectively) and reduced infarction volumes (20.7 +/- 4.5 vs. 12.5 +/- 3.6%, 2,3,5-Triphenyltetrazolium chloride staining) after cerebral ischemia. This observation indicates that the neuroprotection induced by HBO preconditioning may be mediated by an upregulation of HIF-1alpha and its target gene EPO.

Topics: Animals; Behavior, Animal; Blotting, Western; Brain Ischemia; Cerebral Infarction; Cerebrovascular Circulation; DNA; Erythropoietin; Forelimb; Hyperbaric Oxygenation; Hypoxia-Inducible Factor 1, alpha Subunit; Ischemic Preconditioning; Male; Neuroprotective Agents; Oxygen; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tetrazolium Salts; Up-Regulation

Erythropoietin (EPO) has been shown to have an anti-apoptotic action and has the potential to protect against ischaemia/reperfusion injury. This study investigated the effect of high dose EPO (5000 U), administered as a bolus at the onset of reperfusion and at the onset of cold storage in a model of controlled nonheart beating donors kidneys. Porcine kidneys(n = 6) were subjected to 10min warm ischaemia and preserved as follows: Group 1:16 h Cold storage +2 h Normothermic perfusion (16 h CS + 2 h NP) Group 2:16 h CS + 2 h NP + EPO given at the onset of reperfusion Group 3:18 h CS (static hypothermic storage) Group 4:18 h CS + EPO given at the onset of cold storage Haemodynamic and functional parameters were assessed during 3-h reperfusion using autologous blood. Renal blood flow improved in Groups 1 and 2 vs. Groups 3 and 4 though no difference was noted between Groups 3 and 4 (563 +/- 119 vs. 491 +/- 95 vs. 325 +/- 70 vs. 418 +/- 112, respectively; P = 0.012). Total urine output showed no difference between Groups (271 +/- 172 vs. 359 +/- 184 vs. 302 +/- 21 vs. 421 +/- 88; P = 0.576). Percentage serum creatinine fall at 3 h was significantly better in Groups 1 and 2 vs. Group 3 (64 +/- 17 vs. 60 +/- 11 vs. 44 +/- 13 vs. 52 +/- 8; P = 0.04). Fractional-excretion of sodium was significantly lower for Groups 1 and 2 vs. Group 3 and 4 (17 +/- 14 vs. 18 +/- 9 vs. 49 +/- 21 vs. 45 +/- 16 respectively; P = 0.002). There was significant improvement in oxygen consumption in Groups 2 vs. Groups 3 and 4 (P = 0.037) (39 +/- 10 vs. 46 +/- 10 vs. 24 +/- 12 vs. 24 +/- 7 respectively). EPO added at the time of reperfusion improved oxygen consumption when added to NP in comparison to static hypothermic storage but did not exert any other major benefits.

Topics: Animals; Aspartate Aminotransferases; Erythropoietin; Kidney; Kidney Transplantation; Reperfusion Injury; Swine

To investigate the protective effect of erythropoietin (EPO) and its nonhematopoietic derivative (asialoEPO) against intestinal ischemia/reperfusion (I/R) injury in a rat model.. The superior mesenteric artery of Wistar rats was clamped for 60 minutes and then released. The rats were divided into 4 groups (n = 15 in each group): sham operation (Sham), vehicle treatment (Vehicle), EPO treatment (EPO), and asialoEPO treatment (AsialoEPO). EPO and asialoEPO were administered subcutaneously at 1000 units/kg for 10 minutes before clamping, 30 minutes after the start of clamping, and just before declamping. This treatment was followed by determination of 72-hour survival rates, serum TNF-alpha and IL-6 levels, histologic evaluation of the small intestine, quantification of the number of apoptotic cells, and analysis of the antiapoptotic molecules Bcl-xL and XIAP by Western blotting.. The survival rates at 72 hours after I/R injury in the Sham, Vehicle, EPO, and AsialoEPO groups were 100%, 33%, 75%, and 83%, respectively (P < .05). Blood TNF-alpha and IL-6 were significantly more suppressed in the EPO and AsialoEPO groups than in the Vehicle group at 6 hours after I/R injury. Histologically, injury to villi in the EPO and AsialoEPO groups was significantly less than in the Vehicle group. The number of apoptotic cells in the EPO and AsialoEPO groups was significantly less than in the Vehicle group. Western blotting revealed that EPO and asialoEPO constitutively increased the expression of Bcl-xL.. EPO and asialoEPO exert a strong protective effect against intestinal I/R injury, possibly by inhibiting release of TNF-alpha and IL-6 and decreasing apoptosis.

Topics: Animals; Apoptosis; Asialoglycoproteins; Disease Models, Animal; Erythropoietin; Gastrointestinal Agents; Intestinal Diseases; Intestines; Male; Rats; Rats, Wistar; Reperfusion Injury; Survival Analysis

This study was designed to investigate the effects of recombinant erythropoietin (EPO), a hormone widely used for treatment of uremic anemia, in rats subjected to testicular ischemia and reperfusion (I/R).. Thirty-five male rats were divided into the following: control, sham operated, ischemia (I), I/R, and I/R + EPO groups. In the I group, 2 hours of left unilateral testicular torsion were performed, and in the I/R and I/R + EPO groups, an additional 2 hours of testicular detorsions were performed. The I/R + EPO group was pretreated intraperitoneally with EPO (500 IU/kg) before reperfusion. Testicular tissue samples were examined for biochemical and histopathologic parameters. Apoptotic cells in all testes were detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling technique and caspase 3 immunohistochemistry.. At histopathologic examination, ischemic changes in primary spermatocytes were noted in all torted testes. Cellular damage and apoptosis were more severe in ischemic groups than the EPO-pretreated group. There were statistically significant differences in tissue biochemical parameters in the I and I/R groups compared with the I/R + EPO group.. The results of the present study suggest that EPO exerts protective effects against I/R injury via the modulation of free radical scavenger's activities, which decreases lipid peroxidation levels and attenuation of apoptosis.

Topics: Animals; Antioxidants; Apoptosis; Erythropoietin; Germ Cells; Ischemia; Male; Premedication; Protective Agents; Random Allocation; Rats; Recombinant Proteins; Reperfusion; Reperfusion Injury; Spermatic Cord Torsion; Testicular Diseases; Testis

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, partly due to the increased peritubular capillaries. The therapeutic effect of CEPO was evaluated using an endothelial tube formation assay in vitro, and a rat ischemia-reperfusion injury model in vivo. EPO treatment showed the tendency of increased tube formation, while CEPO treatment induced more capillary-like formation than EPO. Ischemia-reperfusion-induced kidneys exhibited characteristic nuclei of apoptosis in tubular epithelial cells with decreased peritubular capillaries, while EPO treatment inhibited tubular apoptosis with preserved endothelial cells. Moreover, CEPO-treated kidneys showed minimal tubular apoptosis with increased peritubular capillary endothelial cells. In conclusion, we identified a new therapeutic approach using CEPO to protect kidneys from ischemia-reperfusion injury by promoting angiogenesis.

Topics: Animals; Capillary Permeability; Cells, Cultured; Coculture Techniques; Endothelium, Vascular; Erythropoietin; Fibroblasts; Humans; Kidney; Male; Neovascularization, Physiologic; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury

We have previously shown that 2 weeks of hypoxia protect the right ventricle of the rat heart from subsequent ischemia and reperfusion (I/R). In the present study, we examined the following: (1) Do shorter periods of hypoxia protect from subsequent I/R? (2) Does intermittent normoxia increase the cardioprotective effect? (3) Is hypoxia-inducible factor-1alpha (HIF-1alpha), erythropoietin (EPO), or vascular endothelial growth factor (VEGF) involved in the protective effects? Preischemic cardiac work was followed by global ischemia, reperfusion, and postischemic cardiac work (15 min each). External heart work was determined at the end of both work phases. Four groups of hearts were investigated: hearts from normoxic rats (n=8), hearts from rats after 24 h of continuous hypoxia (10.5% inspired oxygen, n=7), hearts from rats after 24 h hypoxia with a single intermission of 30 min normoxia (n=9), and hearts from rats after 24 h hypoxia and multiple intermissions of 30 min normoxia (n=7). Protein levels of HIF-1alpha and mRNA levels of EPO and VEGF were determined in right ventricular tissue of normoxic and hypoxic hearts. Postischemic right heart recovery was better in all three hypoxic groups compared with normoxic hearts (61.8 +/- 5.9%, 65.6 +/- 3.0%, and 75.7 +/- 2.6% vs. 46.0 +/- 3.9%, p < 0.01). Hypoxia with multiple normoxic intermissions further improved right heart recovery compared to continuous hypoxia (p < 0.05). HIF-1alpha protein levels were 80.3 +/- 2.5 pg/microg in normoxic hearts and 108.0 +/- 10.3 pg/microg in hypoxic hearts (p = 0.02). No differences in EPO and VEGF mRNA levels were found between normoxic and hypoxic hearts. Twenty-four hours of continuous hypoxia protect the isolated working right heart from subsequent ischemia and reperfusion. When preceding hypoxia is interrupted by multiple reoxygenation periods, there is a further significant increase in cardiac functional recovery. HIF-1alpha may be involved in the protective effect.

Topics: Animals; Erythropoietin; Heart Ventricles; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Ischemic Preconditioning, Myocardial; Male; Oxygen; Rats; Rats, Wistar; Reperfusion Injury; RNA, Messenger; Time Factors; Vascular Endothelial Growth Factor A

To determine whether the administration of erythropoietin at the time of ischaemic renal injury (IRI) inhibits apoptosis, enhances tubular epithelial regeneration and promotes renal functional recovery, as it does in rodent models, in a higher mammalian model.. The model of IRI involved unilateral nephrectomy in pigs, followed a week later by renal artery occlusion for 1 h, followed by reperfusion for 5 days. Pigs were randomized to receive erythropoietin 5000 units/kg intravenously at the time of ischaemia, followed by 1000 units/kg subcutaneously daily, or no treatment (six pigs each). Renal function and structure were analysed; blood and urine were collected daily to determine serum creatinine level, blood urea nitrogen, and creatinine clearance. Animals were killed after 5 days to obtain the injured kidneys. The kidneys were examined histologically for evidence of cellular mitosis, apoptosis and necrosis.. Erythropoietin significantly abrogated renal dysfunction after IRI compared with controls at 12 h after injury; the mean (sem) creatinine clearance (as a percentage of baseline) for IRI was 68.2 (6)% vs erythropoietin-IRI 94.9 (8.9)% (P = 0.027), although by 36 h this was no longer significant, with values of 73.8 (12.7)% vs 95.9 (12)%, respectively (P = 0.23). Erythropoietin also significantly reduced the amount of cell death on histological analysis after 5 days of reperfusion, with a median (range) for IRI of 5.5 (1-45) vs erythropoietin-IRI of 1.5 (0-4) (P = 0.043).. This study confirms the potential clinical applications of erythropoietin as a novel therapeutic agent in patients at risk of IRI.

Topics: Animals; Cell Death; Creatinine; Erythropoietin; Immunohistochemistry; Kidney; Kidney Diseases; Reperfusion Injury; Swine; Treatment Outcome

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.

Topics: Animals; Apoptosis; Cell Proliferation; Erythropoiesis; Erythropoietin; Kidney; Kidney Diseases; Male; Nephritis, Interstitial; Phosphatidylinositol 3-Kinases; Proto-Oncogene Protein c-ets-1; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction

Blood-brain barrier (BBB) leakage plays a role in the pathogenesis of many pathological states of the brain including ischemia and some neurodegenerative disorders. In recent years, erythropoietin (EPO) has been shown to exert neuroprotection in many pathological conditions including ischemia in the brain. This study aimed to investigate the effects of EPO on BBB integrity, infarct size and lipid peroxidation following global brain ischemia/reperfusion in rats. Wistar male rats were divided into four groups (each group n=8); Group I; control group (sham-operated), Group II; ischemia/reperfusion group, Group III; EPO treated group (24 h before decapitation--000 U/kg r-Hu EPO i.p.), Group IV; EPO+ ischemia/reperfusion group (24 h before ischemia/reperfusion--3000 U/kg r-Hu EPO i.p.). Global brain ischemia was produced by the combination of bilateral common carotid arteries occlusion and hemorrhagic hypotension. Macroscopical and spectrophotometrical measurement of Evans Blue (EB) leakage was observed for BBB integrity. Infarct size was calculated based on 2,3,5-triphenyltetrazolium chloride (TTC) staining. Lipid peroxidation in the brain tissue was determined as the concentration of thiobarbituric acid-reactive substances (TBARS) for each group. Ischemic insult caused bilateral and regional BBB breakdown (hippocampus, cortex, corpus striatum, midbrain, brain stem and thalamus). EPO pretreatment reduced BBB disruption, infarct size and lipid peroxide levels in brain tissue with 20 min ischemia and 20 min reperfusion. These results suggest that EPO plays an important role in protecting against brain ischemia/reperfusion through inhibiting lipid peroxidation and decreasing BBB disruption.

Topics: Animals; Blood-Brain Barrier; Brain Infarction; Brain Ischemia; Carotid Arteries; Carotid Stenosis; Disease Models, Animal; Drug Administration Schedule; Epoetin Alfa; Erythropoietin; Evans Blue; Lipid Peroxidation; Male; Neuroprotective Agents; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury; Thiobarbituric Acid Reactive Substances

Erythropoietin (EPO) has been proposed as a novel cytoprotectant in ischemia-reperfusion (I/R) injury of the brain, heart, and kidney. However, whether EPO exerts its protection by prevention of postischemic microcirculatory deterioration is unknown. We have investigated the effect of EPO on I/R-induced microcirculatory dysfunctions. We used the mouse dorsal skinfold chamber preparation to study nutritive microcirculation and leukocyte-endothelial cell interaction in striated muscle of the dorsal skinfold by in vivo fluorescence microscopy before 3 h of ischemia and during 5 days of reperfusion. Animals were pretreated with EPO (5,000 U/kg body wt) 1 or 24 h before ischemia. Vehicle-treated I/R-injured animals served as controls. Additional animals underwent sham operation only or were pretreated with EPO but not subjected to I/R. I/R significantly (P < 0.05) reduced functional capillary density, increased microvascular permeability, and enhanced venular leukocyte-endothelial cell interaction during early reperfusion. These findings were associated with pronounced (P < 0.05) arteriolar constriction and diminution of blood flow during late reperfusion. Pretreatment with EPO induced EPO receptor and endothelial nitric oxide synthase expression at 6 h of reperfusion (P < 0.05). In parallel, EPO significantly (P < 0.05) reduced capillary perfusion failure and microvascular hyperpermeability during early reperfusion and arteriolar constriction and flow during late reperfusion. EPO pretreatment substantially (P < 0.05) diminished I/R-induced leukocytic inflammation by reducing the number of rolling and firmly adhering leukocytes in postcapillary venules. EPO applied 1 h before ischemia induced angiogenic budding and sprouting at 1 and 3 days of reperfusion and formation of new capillary networks at 5 days of reperfusion. Thus our study demonstrates for the first time that EPO effectively attenuates I/R injury by preserving nutritive perfusion, reducing leukocytic inflammation, and inducing new vessel formation.

Topics: Animals; Back; Blood Flow Velocity; Dose-Response Relationship, Drug; Erythropoietin; Mice; Mice, Inbred C57BL; Microcirculation; Muscle, Skeletal; Recombinant Proteins; Reperfusion Injury; Skin

Human recombinant erythropoietin (Epo) has recently been shown to be a potent protector of ischemic damage in various organ systems. A significant reduction of stroke injury following cerebral ischemia has been postulated as well as improved cardiomyocyte function after myocardial infarction in tissue pretreated with Epo. It was the aim of this study to evaluate the effects of Epo in liver ischemia.. Rats were subjected to 45 min of warm hepatic ischemia. Animals were either pretreated with 1,000 IU of Epo in three doses or received 1,000 IU into the portal vein 30 min before ischemia. Control animals received saline at the same time points before ischemia. Animals were than sacrificed 6, 12, 24, 48 h and 7 days after surgery and transaminases were measured. Liver specimens were evaluated regarding apoptosis, necrosis and regeneration capacity.. Apoptosis rates were dramatically reduced in animals pretreated with Epo while mRNA of tumor necrosis factor-alpha and STAT-3 were upregulated in all groups. Intraportal venous injection displayed superiority to subcutaneous preconditioning. Transaminases were significantly reduced among the Epo-treated animals after 6 and 12 h.. Our data suggests a protective effect of Epo in warm hepatic ischemia and reperfusion injury in the rat.

Topics: Animals; Erythropoietin; In Situ Nick-End Labeling; Ischemic Preconditioning; Liver; Male; Rats; Rats, Wistar; Reperfusion Injury; RNA, Messenger; STAT3 Transcription Factor; Tumor Necrosis Factor-alpha; Vascular Endothelial Growth Factor A

Ischemia/reperfusion (I/R) injury is closely associated with tissue damage in various organs, as well as in kidney transplants. Erythropoietin (EPO) has been shown to have a cytoprotective effect against hypoxia. We examined the effect of EPO against renal I/R injury and the underlying mechanism.. Human umbilical vein endothelial cells and human renal proximal tubular epithelial cells were cultured under hypoxic conditions with various EPO concentrations at 37 degrees C and examined the mechanism of cell proliferation by EPO. Moreover, to demonstrate the renoprotective effect in vivo, we treated Sprague-Dawley rats with 100 IU/kg EPO every 2 days for 2 weeks (a total of 6 doses). One day after the last injection, the operations to produce renal I/R injury (bilateral renal occlusion for 60 min) were done, and rats were killed at the end of the reperfusion period (24 hr and 72 hr after reperfusion began).. First, we demonstrated in vitro that EPO increased hypoxia inducible factor-1alpha (HIF-1alpha) expression and stimulated proliferation of both cells under hypoxic conditions. Next, we demonstrated in vivo that EPO treatment increased the number of HIF-1alpha-positive cells, and markedly induced the expression of vascular endothelial growth factor messenger RNA. Using pimonidazole, a molecular probe that detects hypoxia, we found that EPO markedly attenuated tubular hypoxia, and reduced the number of terminal transferase dUTP nick end labeling-positive apoptotic cells and alpha-smooth muscle actin-positive interstitial cells.. We suggested a novel HIF-1alpha induction pathway by EPO under hypoxic conditions. Thus, EPO may protect the kidneys against ischemia reperfusion injury by activating HIF-1alpha.

Topics: Cell Division; Cell Line; Cells, Cultured; Endothelium, Vascular; Erythropoiesis; Erythropoietin; Humans; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Immunohistochemistry; In Situ Nick-End Labeling; Kidney; Reperfusion Injury; RNA, Messenger; Umbilical Veins; Vascular Endothelial Growth Factor A

Testicular torsion is an important clinical urgency. Similar mechanisms occurred after detorsion of the affected testis as in the ischemia reperfusion (I/R) damage. This study was designed to investigate the effects of erythropoietin (EPO) treatment after unilateral testicular torsion. Fifty male Sprague-Dawley rats were divided into five groups. Group 1 underwent a sham operation of the right testis under general anesthesia. Group 2 was same as sham, and EPO (3,000 IU/kg) infused i.p., group 3 underwent a similar operation but the right testis was rotated 720 degrees clockwise for 1 h, maintained by fixing the testis to the scrotum, and saline infused during the procedure. Group 4 underwent similar torsion but EPO was infused half an hour before the detorsion procedure, and in group 5, EPO was infused after detorsion procedure. Four hours after detorsion, ipsilateral and contralateral testes were taken out for evaluation. Treatment with EPO improved testicular structures in the ipsilateral testis but improvement was less in the contralateral testis histologically, but EPO treatment decreased germ cell apoptosis in both testes following testicular IR. TNF-alpha, IL-1beta, IL-6 and nitrite levels decreased after EPO treatment especially in the ipsilateral testis. We conclude that testicular I/R causes an increase in germ cell apoptosis both in the ipsilateral and contralateral testes. Erythropoietin has antiapoptotic and anti-inflammatory effects following testicular torsion.

Topics: Animals; Apoptosis; Disease Models, Animal; Erythropoietin; Hematinics; Inflammation; Male; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Seminiferous Tubules; Spermatic Cord Torsion

The renoprotective effect of erythropoietin (EPO) and the nonhematopoietic EPO, asialoEPO was investigated in a murine ischemia-reperfusion injury (I/R) model.. I/R was created by clamping the right renal pedicle for 60 min after left nephrectomy. Balb/c mice were divided into four groups (n=15 in each group): sham operation (Sham), vehicle treatment (Vehicle), EPO treatment (EPO), and asialoEPO treatment (AsialoEPO). EPO and asialoEPO were given at a dose of 500 IU/kg 30 min before I/R. Plasma creatinine (Cr), survival, and the number of apoptotic cells were analyzed. Protein expression was analyzed by Western blotting.. Plasma Cr level was not significantly different at 6 hr after I/R. At 24 hr after I/R, the Cr (mg/dL) levels in Sham, Vehicle, EPO, and asialoEPO were 0.13+/-0.01, 1.24+/-0.70, 0.24+/-0.08, and 0.25+/-0.13, respectively (P<0.05). The numbers of apoptotic cells in these groups were 0.1+/-0.1, 98.9+/-42.6, 3.3+/-0.7, and 2.9+/-1.6, respectively (P<0.05). Western blotting revealed that in kidney tissue of mice treated with EPO and asialoEPO, p38-MAPK and the proapoptotic molecule Bad was decreased, and the antiapoptotic molecules Bcl-xL and XIAP were increased. Survival rates at 7 days after I/R injury in the Sham, Vehicle, EPO, and AsialoEPO groups were 100%, 21.4%, 23.1%, and 53.8%, respectively (P=0.05).. EPO and asialoEPO attenuated renal dysfunction caused by I/R in mouse kidney at the same level, but only asialoEPO improved survival.

Topics: Animals; Apoptosis; Asialoglycoproteins; bcl-Associated Death Protein; bcl-X Protein; Creatinine; Disease Models, Animal; Erythropoietin; Female; Kidney; Mice; Mice, Inbred BALB C; p38 Mitogen-Activated Protein Kinases; Random Allocation; Reperfusion Injury; Survival Rate; X-Linked Inhibitor of Apoptosis Protein

Testicular torsion is a serious urological emergency, usually involving newborns, children, and adolescents which can lead to subfertility and infertility. Prevention of testicular damage caused by torsion is still a clinical and experimental problem. So far many chemicals and drugs have been investigated for decreasing ischemia/reperfusion (I/R) injury in experimental animals. The possible protective effect of darbepoetin alfa, a novel erythropoietic protein, on testicular tissue after I/R injury was examined in this study.. Thirty rats were divided into three groups: sham operation, torsion/detorsion, and torsion/detorsion plus darbepoetin alfa groups. After torsion (2 hours) and detorsion (4 hours), bilateral orchiectomy was performed. Malondialdehyde, nitric oxide and glutathione levels were determined in testicular tissue.. Administration of darbepoetin alfa caused a decrease of malondialdehyde and nitric oxide levels and an increase in glutathione levels compared with the torsion/detorsion group. In addition, histological injury scores were significantly decreased in the treatment group more than the torsion/detorsion group.. The results suggest that darbepoetin alfa may be a potential protective agent for preventing testicular injury caused by testis torsion.

Topics: Animals; Darbepoetin alfa; Erythropoietin; Hematinics; Male; Rats; Rats, Wistar; Reperfusion Injury; Spermatic Cord Torsion

Recently we found that the level of anti-infarct tolerance afforded by ischemic preconditioning (IPC) and erythropoietin (EPO) infusion was closely correlated with the level of Ser9-phospho-GSK-3beta upon reperfusion in the heart. To get an insight into the mechanism by which phospho-GSK-3beta protects the myocardium from ischemia/reperfusion injury, we examined the effects of IPC and EPO on interactions between GSK-3beta and subunits of the mitochondrial permeability transition pore (mPTP) in this study. Rat hearts were subjected to 25-min global ischemia and 5-min reperfusion in vitro with or without IPC plus EPO infusion (5 units/ml) before ischemia. Ventricular tissues were sampled before or after ischemia/reperfusion to separate subcellular fractions for immunoblotting and immunoprecipitation. Reperfusion increased mitochondrial GSK-3beta by 2-fold and increased phospho-GSK-3beta level in all fractions examined. Major subunits of mPTP, adenine nucleotide translocase (ANT) and voltage-dependent anion channel (VDAC), were co-immunoprecipitated with GSK-3beta after reperfusion. Phospho-GSK-3beta was co-immunoprecipitated with ANT but not with VDAC. IPC+EPO significantly increased the levels of GSK-3beta and phospho-GSK-3beta that were co-immunoprecipitated with ANT to 145+/-8% and 143+/-16%, respectively, of baseline but did not induce phospho-GSK-3beta-VDAC binding. A PKC inhibitor and a PI3 kinase inhibitor suppressed the IPC+EPO-induced increase in the level of phospho-GSK-3beta-ANT complex. The level of cyclophilin D co-immunoprecipitated with ANT after reperfusion was significantly reduced to 39+/-10% of the control by IPC+EPO. These results suggest that reduction in affinity of ANT to cyclophilin D by increased phospho-GSK-3beta binding to ANT may be responsible for suppression of mPTP opening and myocardial protection afforded by IPC+EPO.

Topics: Animals; Disease Models, Animal; Erythropoietin; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; In Vitro Techniques; Intracellular Membranes; Kinetics; Mitochondria, Heart; Mitochondrial Membrane Transport Proteins; Mitochondrial Permeability Transition Pore; Myocardial Ischemia; Phosphorylation; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Ventricular Function

Erythropoietin is traditionally known to regulate erythropoiesis, but recently its protective effect against ischemia-reperfusion injury has been studied in cardiovascular and neuronal systems. This study investigated the effect of recombinant human erythropoietin on ischemia-reperfusion injury in a rat transverse rectus abdominis musculocutaneous (TRAM) flap model.. Twenty-four Sprague-Dawley rats were divided into a control group (n = 12) and a group treated with erythropoietin (n = 12). A superiorly based TRAM flap was elevated and ischemic insult was given for 4 hours. Thirty minutes before reperfusion, single-dose recombinant human erythropoietin (5000 IU/kg) was injected through the intraperitoneal route in the treatment group. At 24 hours postoperatively, systemic neutrophil count, tissue myeloperoxidase activity, amount of malondialdehyde, nitric oxide content, tissue water content, and histologic finding of inflammation were evaluated. At day 10 postoperatively, flap survival rate, angiogenesis, and change in hematocrit level were evaluated.. The myeloperoxidase activity and tissue water content were significantly lower (p < 0.01 and p < 0.005, respectively), and the tissue nitric oxide level was significantly higher (p < 0.005) in the treatment group 24 hours after reperfusion. Perivascular neutrophil infiltration and intravascular adhesion were marked in the control group. Mean flap survival after 10 days was 69 percent in the treatment group and 47 percent in the control group, demonstrating a significant difference (p < 0.005). Neovascularization in the treatment group was also greater than that in the control group. No significant hematocrit rise was noted 10 days after erythropoietin administration.. Recombinant human erythropoietin improved flap survival in ischemia-reperfusion-injured rat TRAM flaps by the possible mechanism of suppressed inflammation, decreased infiltration of neutrophils, increased nitric oxide, and enhanced angiogenesis.

Topics: Animals; Apoptosis; Chemotaxis, Leukocyte; Drug Evaluation, Preclinical; Erythropoietin; Hematocrit; Humans; Inflammation; Malondialdehyde; Neovascularization, Physiologic; Neutrophils; Nitric Oxide; Peroxidase; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Rectus Abdominis; Reperfusion Injury; Surgical Flaps

Erythropoietin (EPO), originally identified for its critical role in promoting erythrocyte survival and differentiation, has been shown to exert multiple paracrine/autocrine functions. Protective effects of EPO have been demonstrated in various tissues and experimental models of ischaemia-induced injury. In the present study, we investigated the effect of EPO on an in vivo rat model of renal ischaemia/reperfusion (I/R) injury and the possible mechanisms implicated in the EPO-mediated anti-apoptotic action.. Male Wistar rats, subjected to renal ischaemia for 45 min, were administered either saline or EPO (500 U/kg, i.p.) 20 min prior to I/R. A sham-operated group served as the control. At 48 h of reperfusion, the renal dysfunction and injury was assessed by measurement of serum biochemical markers (urea, creatinine) and histological grading. Apoptosis was assessed by the TUNEL method and morphological criteria. Expression of Bax and NF-kappaB (p65) was also evaluated.. High levels of serum urea and creatinine were identified at 48 h after ischaemia. The EPO-treated group had significantly lower serum and creatinine levels. Semi-quantitative assessment of the histological lesions showed that rats subjected to I/R developed marked structural damage, whereas significantly less tubular damage was observed in the EPO-treated group. I/R caused an increase in TUNEL-positive cells that was accompanied by morphological evidence of apoptosis. In the EPO-treated rats only a few scattered TUNEL-positive cells were observed. Up-regulation of Bax in the tubular epithelial cells and increased expression of NF-kappaB was observed in the I/R-treated rats, while diminished expression of Bax and positive immunostaining of NF-kappaB was observed in the EPO-treated rats.. Administration of EPO as a single dose before the onset of ischaemia produced a significant reduction in tubular injury, which was accompanied by a marked amelioration of renal functional impairment. The cytoprotective action of EPO against I/R injury seems to be associated with its anti-apoptotic action. Moreover, transcription factor NF-kappaB is likely to play a pivotal role in the pathophysiology of I/R renal injury and might have a key role in EPO-mediated protective effects.

Topics: Acute Kidney Injury; Animals; Apoptosis; Disease Models, Animal; Erythropoietin; Male; NF-kappa B; Rats; Rats, Wistar; Reperfusion Injury

Several previous studies have demonstrated a beneficial effect of the adenosine receptor (AdoR) antagonist theophylline in different forms of acute renal failure in laboratory animals and in humans. Therefore, we wanted to test whether theophylline can also improve impaired allograft function following ischemia reperfusion injury in experimental kidney transplantation (KT). Orthotopic transplantation of the left kidney was performed from Fisher 344 into Lewis rats. All transplanted rats received daily cyclosporine (5 mg/kg). The effect of theophylline treatment (10 mg/kg) on graft function was compared with appropriate controls on day 5 after KT by assessment of glomerular filtration rate (GFR) (inulin clearance). On day 5, GFR of allografts in control rats was 0.23 +/- 0.05 ml/min/g kidney weight (n = 10) compared with 0.50 +/- 0.09 ml/min/g in rats receiving theophylline (n = 9, p < 0.01), representing a 2-fold increase in GFR. Renal AdoR A(1) mRNA content was significantly increased in both KT groups compared with their respective control groups, whereas mRNA of AdoR A(2a), A(2b), and A(3) were found to be unchanged. Theophylline did not affect significantly interstitial infiltration of the graft by monocytes/macrophages and T-cells. Likewise, serum cytokines [interleukin (IL)-2, IL-6, IL-10, tumor necrosis factor-alpha] and erythropoietin plasma levels were not different among the allograft groups. The present study demonstrates that theophylline remarkably improved early renal allograft function in rats undergoing KT without influencing cytokine serum patterns or tissue inflammation. Since theophylline is a commonly used medication in humans, clinical studies in patients undergoing KT are warranted.

Topics: Animals; Cytokines; Disease Models, Animal; Erythropoietin; Glomerular Filtration Rate; Immunohistochemistry; Kidney; Kidney Cortex; Kidney Transplantation; Male; Purinergic P1 Receptor Antagonists; Rats; Rats, Inbred F344; Rats, Inbred Lew; Receptors, Purinergic P1; Reperfusion Injury; Theophylline

This study was designed to investigate the neuroprotective effect of intrinsic and extrinsic erythropoietin (EPO) against hypoxia/ischemia, and determine the optimal time-window with respect to the EPO-induced neuroprotection. Experiments were conducted using primary mixed neuronal/astrocytic cultures and neuron-rich cultures. Hypoxia (2%) induces hypoxia-inducible factor-1alpha (HIF-1alpha) activity followed by strong EPO expression in mixed cultures and weak expression in neuron-rich cultures as documented by both western blot and RT-PCR. Immunoreactive EPO was strongly detected in astrocytes, whereas EPOR was only detected in neurons. Neurons were significantly damaged in neuron-rich cultures but were distinctly rescued in mixed cultures. Application of recombinant human EPO (rhEPO) (0.1 U/mL) within 6 h before or after hypoxia significantly increased neuronal survival compared with no rhEPO treatment. Application of rhEPO after onset of reoxygenation achieved the maximal neuronal protection against ischemia/reperfusion injury (6 h hypoxia followed 24 h reoxygenation). Our results indicate that HIF-1alpha induces EPO gene released by astrocytes and acts as an essential mediator of neuroprotection, prove the protective role of intrinsic astrocytic-neuronal signaling pathway in hypoxic/ischemic injury and demonstrate an optimal therapeutic time-window of extrinsic rhEPO in ischemia/reperfusion injury in vitro. The results point to the potential beneficial effects of HIF-1alpha and EPO for the possible treatment of stroke.

Topics: Animals; Brain Ischemia; Cells, Cultured; Cerebral Cortex; Erythropoietin; Humans; Neurons; Neuroprotective Agents; Rats; Rats, Wistar; Receptors, Erythropoietin; Recombinant Proteins; Reperfusion Injury

Laparoscopic surgery has gained wide acceptance for almost every kind of surgical procedure, although it has produced significant oxidative injury to intraabdominal organs depending on the pressure level and the kind of the gas used. The literature describes several preventive measures for decreasing the postlaparoscopic oxidative injury such as low intraabdominal pressure, gasless laparoscopy, and laparoscopic preconditioning. Erythropoietin was shown previously to decrease ischemia-reperfusion injury to the liver. The current study evaluated the effect of erythropoietin against laparoscopy-induced oxidative injury, as compared with laparoscopic preconditioning.. For this study, 64 male Spraque-Dawley rats were randomly assigned to one of the following groups. The control group was subjected to a sham operation. The laparoscopy group was subjected to 60 min of pneumoperitoneum. The laparoscopic preconditioning plus laparoscopy group was subjected to 5 min of insufflation and 5 min of desufflation followed by 60 min of pneumoperitoneum. The erythropoietin plus laparoscopy group was subjected to a subcutaneous injection of erythropoietin as a single 1,000-U/kg dose followed by 60 min of pneumoperitoneum. After 45 min of desufflation subsequent to cessation of pneumoperitoneum, blood, liver, and kidney samples were obtained from half of the rats. The other half of the rats were observed for a reperfusion period of 24 h. Tissue and blood samples also were obtained after this period.. Laparoscopy produced significant oxidative injury, as compared with the sham treatment. Laparoscopic preconditioning produced significant amelioration of the ischemic injury. Although erythropoietin administration during the prelaparoscopic period decreased the pneumoperitoneum-induced oxidative injury, the beneficial effect of laparoscopic preconditioning was more pronounced.. Laparoscopic preconditioning is more effective than the preischemic administration of erythropoietin in reducing laparoscopy-induced oxidative injury.

Topics: Animals; Erythropoietin; Injections, Subcutaneous; Insufflation; L-Lactate Dehydrogenase; Laparoscopy; Male; Malondialdehyde; Oxidative Stress; Pneumoperitoneum, Artificial; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Tumor Necrosis Factor-alpha

Administration of human recombinant erythropoietin (EPO) at time of acute ischemic renal injury (IRI) inhibits apoptosis, enhances tubular epithelial regeneration, and promotes renal functional recovery. The present study aimed to determine whether darbepoetin-alfa (DPO) exhibits comparable renoprotection to that afforded by EPO, whether pro or antiapoptotic Bcl-2 proteins are involved, and whether delayed administration of EPO or DPO 6 h following IRI ameliorates renal dysfunction. The model of IRI involved bilateral renal artery occlusion for 45 min in rats (N = 4 per group), followed by reperfusion for 1-7 days. Controls were sham-operated. Rats were treated at time of ischemia or sham operation (T0), or post-treated (6 h after the onset of reperfusion, T6) with EPO (5000 IU/kg), DPO (25 mug/kg), or appropriate vehicle by intraperitoneal injection. Renal function, structure, and immunohistochemistry for Bcl-2, Bcl-XL, and Bax were analyzed. DPO or EPO at T0 significantly abrogated renal dysfunction in IRI animals (serum creatinine for IRI 0.17 +/- 0.05 mmol/l vs DPO-IRI 0.08 +/- 0.03 mmol/l vs EPO-IRI 0.04 +/- 0.01 mmol/l, P = 0.01). Delayed administration of DPO or EPO (T6) also significantly abrogated subsequent renal dysfunction (serum creatinine for IRI 0.17 +/- 0.05 mmol/l vs DPO-IRI 0.06 +/- 0.01 mmol/l vs EPO-IRI 0.03 +/- 0.03 mmol/l, P = 0.01). There was also significantly decreased tissue injury (apoptosis, P < 0.05), decreased proapoptotic Bax, and increased regenerative capacity, especially in the outer stripe of the outer medulla, with DPO or EPO at T0 or T6. These results reaffirm the potential clinical application of DPO and EPO as novel renoprotective agents for patients at risk of ischemic acute renal failure or after having sustained an ischemic renal insult.

Topics: Acute Kidney Injury; Animals; Apoptosis; bcl-2-Associated X Protein; bcl-X Protein; Cell Division; Creatinine; Darbepoetin alfa; Disease Models, Animal; Erythropoietin; Immunohistochemistry; Injections, Intraperitoneal; Kidney Tubular Necrosis, Acute; Kidney Tubules, Proximal; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Regeneration; Reperfusion Injury; Time Factors

Based on the findings that erythropoietin (EPO) has been proved to be a multiple functional cytokine to attenuate ischemia-reperfusion (I/R) injury in various organs such as brain, heart, and kidney in animals, this experiment was designed to evaluate the effect of pretreatment with recombined human erythropoietin (rhEPO) on I/R-induced lung injury.. Left lungs of rats underwent 90 min of ischemia and then were reperfused for up to 2 h. Animals were randomly divided into three experimental groups as sham group, I/R group, and rhEPO + I/R group (a single dose of rhEPO was injected intraperitoneally 3000 U/kg 24 h prior to operation). Lung injury was evaluated according to semi-quantitative analysis of microscopic changes, tissue polymorphonuclear neutrophils (PMNs) accumulation (myeloperoxidase (MPO) activity), and pulmonary microvascular permeability (Evan's blue dying method). Peripheral arterial and venous blood samples were obtained for blood-gas analysis after 5 min occlusion of right lung hilus at the end of reperfusion. The serum concentration of tumor necrosis factor (TNF)-alpha was also measured by the method of enzyme-linked immunosorbent assay.. Histological injury scoring revealed significantly lessened lung alveolus edema and neutrophils infiltration in the rhEPO pretreated group compared with I/R group (p < 0.05). The rhEPO pretreated animals exhibited markedly decreased lung microvascular permeability (p < 0.05) and myeloperoxidase activity (p < 0.05). Blood-gas analysis demonstrated that the pretreated animals had significantly ameliorated pulmonary oxygenation function (p < 0.05). The serum concentration of tumor necrosis factor-alpha in rhEPO pretreated group was markedly decreased compared with that of I/R group (p < 0.05).. Pretreatment with rhEPO appears to attenuate I/R-induced lung injury. This function is partly related with the capacity that rhEPO inhibits the accumulation of polymorphonuclear neutrophils in lung tissue and decreases the systematic expression of tumor necrosis factor-alpha.

Topics: Animals; Capillary Permeability; Erythropoietin; Lung; Male; Oxygen; Partial Pressure; Peroxidase; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Respiratory Distress Syndrome; Tumor Necrosis Factor-alpha

Topics: Cardiotonic Agents; Dose-Response Relationship, Drug; Drug Administration Schedule; Erythropoietin; Humans; Myocardial Ischemia; Neovascularization, Physiologic; Reperfusion Injury; Ventricular Remodeling

Recently, erythropoietin was shown to have both hematopoietic as well as tissue-protective properties. Erythropoietin (EPO) had a protective effect in animal models of cerebral ischemia, mechanical trauma of the nervous system, myocardial infarction, and ischemia-reperfusion (I/R) injury of the kidney. It is not known whether EPO protects the liver against I/R injury. Using a rat model of liver I/R injury, we aimed to determine the effect of the administration of human recombinant erythropoietin (rhEPO) on liver injury. Rats were subjected to 30 min of liver ischemia followed by 2 h of reperfusion. When compared with the sham-operated rats, I/R resulted in significant rises in the serum levels of aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, gamma-glutamyl transferase, tissue lipid peroxidation, caspase-3 activity and altered histology. Administration of rhEPO 5 min before ischemia was able to reduce the biochemical evidence of liver injury; however, this protection was not evident when rhEPO was administered 5 min before reperfusion. Mechanistically, early administration of rhEPO was able to reduce the oxidative stress and caspase-3 activation, suggesting the subsequent reduction of apoptosis. This study provides the first evidence that rhEPO causes a substantial reduction of the liver injury induced by I/R in the rat.

Topics: Animals; Biochemistry; Caspases; Erythropoietin; Humans; Ischemia; Liver; Male; Malondialdehyde; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury; Time Factors

To examine the changes in erythropoietin (Epo) protein and its mRNA expression in rat brain subjected to focal ischemia and possible mechanism of the preconditioning of mitochondrial toxin 3-nitropropionic acid (3-NPA), rats were administrated either vehicle or 3-NPA at a dose of 20 mg/kg, intraperitoneally (ip), 3 days prior to a 2-h middle cerebral artery occlusion followed by 24-h reperfusion. Infarct volumes were measured by using 2, 3, 5 triphenylte trazolinm chloride (TTC) staining, and Epo protein and its mRNA levels were assessed by immunohistochemistry and reverse transcriptase polymerase chain reaction (RT-PCR), respectively. Our results showed that after reperfusion, Epo was found to be expressed extensively in the rat brain. It was most apparent in the basal nuclei and hippocampus, and was, to some extent, present in cortex. Preconditioning with 3-NPA caused a reduction in infarct volume. The expression of both Epo protein and mRNA increased significantly in the different brain areas in the 3-NPA pretreated group as compared with the non-pretreated ischemia model group. These results suggested that preconditioning with low dose 3-NPA could induce ischemic tolerance and neuro-protective effects by increasing the Epo expression in the ischemic and ischemia-related areas.

Topics: Animals; Erythropoietin; Immunohistochemistry; Infarction, Middle Cerebral Artery; Ischemic Preconditioning; Male; Nitro Compounds; Propionates; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

Neonatal stroke is a condition that leads to disability in later life, and as yet there is no effective treatment. Recently, erythropoietin (EPO) has been shown to be cytoprotective following brain injury and may promote neurogenesis. However, the effect of EPO on functional outcome and on morphologic changes in neonatal subventricular zone (SVZ) following experimental neonatal stroke has not been described. We used a transient focal model of neonatal stroke in P10 rat. Injury was documented by diffusion weighted MRI during occlusion. Immediately upon reperfusion, either EPO (5U/gm) or vehicle was administered intraperitoneally and animals were allowed to grow for 2 wk. Sensorimotor function was assessed using the cylinder rearing test and then brains were processed for volumetric analysis of the SVZ. Stroke induced SVZ expansion proportional to hemispheric volume loss. EPO treatment markedly preserved hemispheric volume and decreased the expansion of SVZ unilaterally. Furthermore, EPO treatment significantly improved the asymmetry of forelimb use following neonatal stroke. This functional improvement directly correlated with the amount of preserved hemispheric volume. These results suggest EPO may be a candidate in the treatment of neonatal stroke.

Topics: Animals; Animals, Newborn; Behavior, Animal; Benzoxazines; Brain; Brain Injuries; Coloring Agents; Diffusion; Erythropoietin; Hypoxia-Ischemia, Brain; Magnetic Resonance Imaging; Neurons; Oxazines; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Reperfusion Injury; Stroke; Time Factors; Treatment Outcome

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

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

Hypoxia-inducible factor-1 (HIF-1) is a key regulator of the cellular hypoxic response. We previously showed that HIF-1 activation is essential for heat acclimation (AC) in Caenorhabditis elegans. Metabolic changes in AC rat hearts indicate HIF-1alpha activation in mammals as well. Here we characterize the HIF-1alpha profile and the transcriptional activation of its target genes following AC and following heat stress (HS) in hearts from nonacclimated (C; 24 degrees C) and AC (34 degrees C, 1 mo) rats. We used Western blot and immunohistochemistry to measure HIF-1alpha levels and EMSA and RT-PCR/quantitative RT-PCR to detect expression of the HIF-1alpha-targeted genes, including vascular endothelial growth factor (Vegf), heme oxygenase-1 (HO1), erythropoietin (Epo), and Epo receptor (EpoR). EpoR and Epo mRNA levels were measured to determine systemic effects in the kidneys and cross-tolerance effects in C and AC ischemic hearts (Langendorff, 75% ischemia, 40 min). The results demonstrated that 1) after AC, HIF-1alpha protein levels were increased, 2) HS alone induced transient HIF-1alpha upregulation, and 3) VEGF and HO1 mRNA levels increased after HS, with greater magnitude in the AC hearts. Epo mRNA in AC kidneys and EpoR mRNA in AC hearts were also elevated. In AC hearts, EpoR expression was markedly higher after HS or ischemia. Hearts from AC rats were dramatically protected against infarction after ischemia-perfusion. We conclude that HIF-1 contributes to the acclimation-ischemia cross-tolerance mechanism in the heart by induction of both chronic and inducible adaptive components.

Topics: Animals; Blotting, Western; Body Temperature; Caenorhabditis elegans; Cell Nucleus; Cytosol; Erythropoietin; Heart; Heme Oxygenase-1; Hot Temperature; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Immunohistochemistry; Immunoprecipitation; Ischemia; Kidney; Male; Microscopy, Confocal; Microscopy, Fluorescence; Models, Statistical; Myocardium; Rats; Receptors, Erythropoietin; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Subcellular Fractions; Temperature; Time Factors; Transcription, Genetic; Transcriptional Activation; Up-Regulation; Vascular Endothelial Growth Factor A

We previously demonstrated that chronic hypoxia has pivotal roles in the progression of tubulointerstitial injury from the early stage of the uninephrectomized Thy1 nephritis model. We have also shown that pretreatment of cobalt confers renoprotection in the ischemia/reperfusion (I/R) injury, in association with the up-regulation of hypoxia-inducible factor (HIF)-regulated genes. Here, we tested the hypothesis that cobalt administration not only attenuates acute ischemic insult, but also ameliorates tubulointerstitial injury secondary to chronic hypoxia.. We applied sustained cobalt treatment to the uninephrectomized Thy1 nephritis model at 3 to 5 weeks, when tubular hypoxia appeared. Histologic evaluation, including glomerular and peritubular capillary networks, was made at 8 weeks. HIF activation was confirmed by real-time polymerase chain reaction (PCR) analyses for HIF-regulated genes, such as erythropoietin (EPO), vascular endothelial growth factor (VEGF), and heme oxygenase 1 (HO-1). Up-regulation of HIF-1alpha and HIF-regulated genes was also verified by Western blotting analysis. To elucidate responsible mechanisms of cobalt in the amelioration of tubuloniterstitial injury, terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) nick end labeling (TUNEL) staining was conducted at 5 weeks. A combination therapy with angiotensin receptor blocker (ARB), olmesartan, was also challenged.. Although the intervention did not change glomerular structural damage or urinary protein excretion rate, tubulointerstitial injury was improved in cobalt-treated animals when compared with the vehicle-treated group. The amelioration was associated with the parallel up-regulation of renoprotective, HIF-regulated gene expression. TUNEL staining revealed that the number of apoptotic cells was reduced in the cortex by cobalt administration, suggesting that renoprotection was achieved partly through its antiapoptotic properties. Furthermore, it was demonstrated that cobalt treatment exerts additional renoprotective effects with the ARB treatment in this model.. Maneuvers to activate HIF in the ischemic tubulointerstitium will be a new direction to future therapeutic strategies.

Topics: Animals; Antibodies; Antimutagenic Agents; Apoptosis; Capillaries; Cobalt; Erythropoietin; Gene Expression Regulation; Glomerulonephritis; Glomerulosclerosis, Focal Segmental; Heme Oxygenase-1; Hypoxia; Hypoxia-Inducible Factor 1; In Situ Nick-End Labeling; Isoantibodies; Kidney Tubules; Male; Nephrectomy; Rats; Rats, Sprague-Dawley; Renin-Angiotensin System; Reperfusion Injury; Vascular Endothelial Growth Factor A

Multiple protective effects of erythropoietin (EPO), such as antiapoptotic, antioxidant, angiogenic and neuroprotective effects, against ischemia have been demonstrated in cell culture and animal models. Genistein is also a potent tyrosine kinase inhibitor. The aims of the present study were to evaluate the effects of EPO on renal ischemia/reperfusion injury and to determine the role of the tyrosine kinase pathway on this process.. Sprague-Dawley rats were assigned to five groups: (i) sham (Group I); (ii) control with renal ischemia (right nephrectomy and clamping on the left renal pedicle for 45 min and reperfusion; Group II); (iii) EPO + ischemia (Group III); (iv) genistein (an inhibitor of tyrosine kinase) + ischemia (Group IV); and (v) EPO + genistein + ischemia (Group V). Recombinant human EPO (1000 IU/kg) and genistein (10 mg/kg) were given 2 hours before ischemia. Blood samples and the left kidney were obtained after 45 min of reperfusion from half of the rats and after 24 h from the other half.. The blood urea nitrogen, creatinine, tumour necrosis factor-alpha (P < 0.05) and interleukin-2 (P < 0.01) levels, and renal tissue lipid peroxidation (P < 0.05) were significantly lower in Group III than in Group II at 45 min of reperfusion. Following 24 h of reperfusion, EPO decreased tissue peroxidation and histopathological injury, whereas genistein reversed it. The most prominent ischemic injury was observed in Group IV in which genistein was administered. There was no significant difference between Groups II and V.. These results suggest that EPO is effective in attenuating renal ischemia/reperfusion injury, and this effect may be related to tyrosine kinase activity.

Topics: Animals; Enzyme Inhibitors; Erythropoietin; Genistein; Interleukin-2; L-Lactate Dehydrogenase; Lipid Peroxidation; Male; Protein-Tyrosine Kinases; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Tumor Necrosis Factor-alpha

To investigate the protective effect of recombinant human erythropoietin (rhEPO) on retinal ischemic injury induced by raising intraocular pressure.. Twelve New Zealand rabbits were divided into the model group (n=6) and the EPO group (n=6). Either of bilateral eyes was randomly made into the acutely high intraocular pressure model using the method of saline perfusion into anterior chamber. After the model was made, rhEPO 100 IU x kg(-1) was injected into the hypodermic tissue of EPO group rabbits twice a week for one week. Flash electroretinogram (ERG) was detected 30 min before and at 1, 3, 7, and 14 days after perfusion, respectively.. The amplitude of ERG-b wave had no significant difference in the EPO group compared with that in the model group before anterior chamber perfusion (P > 0.05). The amplitude of ERG-b wave dropped down to the lowest at 1 day after perfusion and could not come back to the baseline in the model group (P < 0.05). The resemble situation was in the EPO group except that the amplitude of ERG-b wave came back to the baseline at 14 days (P > 0.05).. EPO can improve the amplitude of ERG-b wave following retinal ischemia. This indicates that EPO has the potential to be an optimal neuroprotective agent.

Topics: Animals; Electroretinography; Erythropoietin; Female; Humans; Male; Neuroprotective Agents; Ocular Hypertension; Rabbits; Recombinant Proteins; Reperfusion Injury; Retina; Retinal Diseases

Erythropoietin (EPO), the principal hematopoietic cytokine that regulates mammalian erythropoiesis, exhibits diverse cellular effects in non-hematopoietic tissues. The physiologic functions of EPO are mediated by its specific cell-surface receptor EPOR. In this study, we demonstrate EPOR expression in adult rat cardiac myocytes and examine the direct effects of EPO on the heart to investigate whether recombinant EPO may exert an acute cardioprotective effect during ischemia-reperfusion injury. To determine whether EPO is cardioprotective, isolated rat hearts were perfused for 10 min in the Langendorff-mode with Krebs-Henseleit buffer in the absence or presence of brief recombinant EPO treatment while left-ventricular-developed pressure (LVDP) was measured continuously to assess contractile function. The hearts were then subjected to 20 min of normothermic global ischemia followed by 25 min of reperfusion. The post-ischemic recovery of LVDP in the untreated control hearts was 26 +/- 5% of their baseline LVDP, whereas hearts pretreated with EPO exhibited significantly improved post-ischemic recovery to 57 +/- 7%. We used 31P nuclear magnetic resonance (NMR) spectroscopy to determine whether modulation of intracellular pH and/or high-energy phosphate levels during ischemia contributed to EPO-mediated cardioprotection. These experiments revealed that the rapid cardioprotective effect of EPO during ischemia-reperfusion injury was associated with preservation of ATP levels in the ischemic myocardium.

Topics: Adenosine Triphosphate; Animals; Buffers; Cardiotonic Agents; Erythropoietin; Heart; Hemodynamics; Hydrogen-Ion Concentration; Magnetic Resonance Spectroscopy; Male; Myocardial Contraction; Myocardium; Myocytes, Cardiac; Perfusion; Phosphocreatine; Rats; Rats, Sprague-Dawley; Receptors, Erythropoietin; Recombinant Proteins; Reperfusion Injury; RNA, Messenger; Time Factors

Erythropoietin is protective against cardiac ischemia, but the underlying mechanisms are unknown. We determined whether erythropoietin (0.5 - 10.0 U/ml) confers acute cardioprotection in infant rabbit hearts and the contribution of protein kinases, nitric oxide synthase and potassium channels to the underlying mechanism. Hearts from normoxic infant New Zealand White rabbits (n=8/group) were isolated and perfused in the Langendorff mode. Biventricular function was recorded under steady-state conditions prior to 30 min global no-flow ischemia and 35 min reperfusion. Administration of erythropoietin for 15 min immediately prior to ischemia resulted in a concentration-dependent increase in recovery of left and right ventricular developed pressure in rabbit hearts following myocardial ischemia and reperfusion. The optimal concentration of erythropoietin that afforded maximum recovery of developed pressure was manifest at 1.0 U/ml. Erythropoietin (1.0 U/ml) treatment resulted in phosphorylation of PKC, p38 MAP kinase and p42/44 MAP kinase. The cardioprotective effects of erythropoietin were abolished by the protein kinase inhibitors SB203580 (p38 MAP kinase), PD98059 (p42/44 MAP kinase) and chelerythrine (PKC) as well as the potassium channel blockers glibenclamide, HMR 1098, 5-HD and Paxilline. Nitrite and nitrate release from hearts before (2.3 +/- 0.9 nmol/min/g) and after (2.4 +/- 1.9 nmol/min/g) 15 min treatment with erythropoietin (1.0 U/ml) were not different. L-NAME and L-NMA did not block the cardioprotective effect of erythropoietin. We conclude the rapid activation of potassium channels and protein kinases by erythropoietin represents an important new mechanism for increasing cardioprotection.

Topics: Animals; Blotting, Western; Dose-Response Relationship, Drug; Erythropoietin; Heart; Myocardial Ischemia; Nitric Oxide Synthase; Organ Culture Techniques; Potassium Channels; Protein Kinase Inhibitors; Protein Kinases; Rabbits; Reperfusion Injury; Signal Transduction

Here we investigate the effects of erythropoietin (EPO) on the tissue/organ injury caused by hemorrhagic shock (HS), endotoxic shock, and regional myocardial ischemia and reperfusion in anesthetized rats. Male Wistar rats were anesthetized with thiopental sodium (85 mg/kg i.p.) and subjected to hemorrhagic shock (HS; i.e., mean arterial blood pressure reduced to 45 mmHg for 90 min, followed by resuscitation with shed blood for 4 h), endotoxemia (for 6 h), or left anterior descending coronary artery occlusion (25 min) and reperfusion (2 h). HS and endotoxemia resulted in renal dysfunction and liver injury. Administration of EPO (300 IU/kg i.v., n = 10) before resuscitation abolished the renal dysfunction and liver injury in hemorrhagic, but not endotoxic, shock. HS also resulted in significant increases in the kidney of the activities of caspases 3, 8, and 9. This increase in caspase activity was not seen in HS rats treated with EPO. In cultured human proximal tubule cells, EPO concentration-dependently reduced the cell death and increase in caspase-3 activity caused by either ATP depletion (simulated ischemia) or hydrogen peroxide (oxidative stress). In the heart, administration of EPO (300 IU/kg i.v., n = 10) before reperfusion also caused a significant reduction in infarct size. In cultured rat cardiac myoblasts (H9C2 cells), EPO also reduced the increase in DNA fragmentation caused by either serum deprivation (simulated ischemia) or hydrogen peroxide (oxidative stress). We propose that the acute administration of EPO on reperfusion and/or resuscitation will reduce the tissue injury caused by ischemia-reperfusion of the heart (and other organs) and hemorrhagic shock.

Topics: Animals; Blood Pressure; Disease Models, Animal; Erythropoietin; Heart Rate; Male; Myocardial Ischemia; Rats; Reperfusion Injury; Shock, Hemorrhagic

Ischemia-induced acute renal failure (ARF) is known to be associated with significant impairment of urinary concentrating ability and down-regulation of renal aquaporins (AQPs) and sodium transporters in rats. We tested whether treatment with erythropoietin (EPO) or alpha-melanocyte-stimulating hormone (alpha-MSH) in combination with EPO reduces the renal ischemia/reperfusion (I/R) injury and prevents the down-regulation of renal AQPs and major sodium transporters.. I/R-induced ARF was established in rats by 40-minute temporary bilateral obstruction of renal arteries, and rats were kept in metabolic cages for urine measurements. After 2 or 4 days following EPO and/or alpha-MSH treatment, kidneys were removed to determine the expression levels of AQPs and sodium transporters by semiquantitative immunoblotting.. Rats with ARF showed significant renal insufficiency, increased urine output, and high fractional excretion of urinary sodium. Consistent with this, immunoblotting and immunocytochemistry revealed that the kidney expression of AQPs (AQP-1, -2 and -3) and sodium transporters [Na,K-ATPase, rat type 1 bumetanide-sensitive Na-K-2Cl cotransporter (BSC-1), Na/H exchanger type 3 (NHE3), and thiazide-sensitive sodium chloride cotransporter (TSC)] in ARF rats was significantly decreased compared to sham-operated control rats. In contrast, EPO treatment at the time of ischemia of rats with ARF significantly prevented the ischemia-induced down-regulation of renal AQPs and sodium transporters and in parallel improved the urinary concentrating capability and renal sodium reabsorption. Importantly, similar effects were observed following the initiation of EPO or alpha-MSH treatment 4 hours after the onset of ischemia injury. Moreover, the combination of EPO with alpha-MSH potentiated the beneficial effects of single compound treatment.. EPO and/or alpha-MSH treatment significantly prevent I/R-induced injuries such as urinary-concentrating defects and down-regulation of renal AQPs and sodium transporters.

Topics: Acute Kidney Injury; alpha-MSH; Animals; Antibody Specificity; Aquaporin 1; Aquaporin 2; Aquaporin 3; Aquaporins; Down-Regulation; Erythropoietin; Immunoblotting; Kidney; Male; Rats; Rats, Wistar; Reperfusion Injury; Sodium; Sodium-Hydrogen Exchanger 3; Sodium-Hydrogen Exchangers; Sodium-Potassium-Exchanging ATPase; Water

Erythropoietin (EPO) is upregulated by hypoxia and causes proliferation and differentiation of erythroid progenitors in the bone marrow through inhibition of apoptosis. EPO receptors are expressed in many tissues, including the kidney. Here it is shown that a single systemic administration of EPO either preischemia or just before reperfusion prevents ischemia-reperfusion injury in the rat kidney. Specifically, EPO (300 U/kg) reduced glomerular dysfunction and tubular injury (biochemical and histologic assessment) and prevented caspase-3, -8, and -9 activation in vivo and reduced apoptotic cell death. In human (HK-2) proximal tubule epithelial cells, EPO attenuated cell death in response to oxidative stress and serum starvation. EPO reduced DNA fragmentation and prevented caspase-3 activation, with upregulation of Bcl-X(L) and XIAP. The antiapoptotic effects of EPO were dependent on JAK2 signaling and the phosphorylation of Akt by phosphatidylinositol 3-kinase. These findings may have major implications in the treatment of acute renal tubular damage.

Topics: Animals; Apoptosis; Blood Proteins; Caspase 3; Caspase Inhibitors; Caspases; Cell Line, Transformed; Creatinine; Disease Models, Animal; Erythropoietin; Humans; Kidney Diseases; Kidney Tubules; Male; Oxidative Stress; Phosphorylation; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Rats; Rats, Wistar; Reperfusion Injury

Erythropoietin (EPO) is a potent stimulator of erythroid progenitor cells and is known to be up-regulated during states of hypoxia. Here we investigate the effects of renal ischemia/reperfusion (I/R) on the degree of renal dysfunction and injury with recombinant human EPO in mice when given as either a 3-day pretreatment, or upon reperfusion of the kidney.. Mice were treated with EPO (1000 IU/kg/day subcutaneously) for 3 days, or treated with EPO (1000 IU/kg subcutaneously) upon reperfusion, and subsequently subjected to bilateral renal artery occlusion (30 minutes) and reperfusion (24 hours). At the end of experiments, the following indicators and markers of renal injury and dysfunction were measured: plasma urea, creatinine, and aspartate aminotransferase (AST), tissue myeloperoxidase (MPO) activity [for polymorphonuclear leukocyte (PMN) infiltration], and tissue malonaldehyde (MDA) levels (for tissue lipid peroxidation). Kidneys were used for histologic evaluation of renal injury.. EPO was able to significantly attenuate the renal dysfunction and injury associated with I/R, as well as the tissue injury. The increase in renal MPO activity and, hence, the degree of PMN infiltration were also significantly reduced in EPO-treated mice. In addition, lipid peroxidation as a result of renal I/R injury was also attenuated in EPO-treated mice.. The protection afforded by the pretreatment regime of EPO was greater than that of administering EPO as a single bolus upon reperfusion. We propose that different mechanisms underlie the protective effects seen with EPO when given as either a daily pretreatment or as a single bolus, which need to be further investigated.

Topics: Animals; Aspartate Aminotransferases; Creatinine; Erythropoietin; Male; Mice; Mice, Inbred C57BL; Nephritis; Reperfusion Injury; Urea

Erythropoietin (EPO) is a hormone known to stimulate hematopoiesis. However, recent research suggests additional properties of EPO, such as protection against ischemia/reperfusion (I/R) injury in various tissues. We studied the effect of timing of EPO administration on cardioprotection during I/R in the heart. Male Sprague-Dawley rats were subjected to 45 minutes of coronary occlusion, followed by 24 hours of reperfusion. Animals were randomized to receive saline or single dose of EPO (5,000 IU/kg) either 2 hours before I/R, at the start of ischemia, or after the onset of reperfusion. The ratio of infarct area/area at risk (planimetry), left ventricular (LV) function (pressure development), and apoptosis (number of active caspase-3 positive cells) were determined after 24-hour reperfusion. Administration of EPO during different time points resulted in a 19 to 23% (P < 0.05) reduction in the infarct area/area at risk, which was accompanied by a trend toward better LV hemodynamic parameters. Apoptosis was significantly attenuated in groups treated with EPO at the start of ischemia (29% reduction) and after the onset of reperfusion (38%), and to a lesser extent (16%) in the group pre-treated with EPO. Thus, in vivo administration of EPO at different time points protects the myocardial structure and preserves cardiac function during I/R. Cardioprotective effect of EPO is associated with inhibition of apoptosis.

Topics: Animals; Apoptosis; Cardiotonic Agents; Cell Proliferation; Drug Administration Schedule; Erythropoietin; Hemodynamics; Ki-67 Antigen; Male; Myocardial Infarction; Myocytes, Cardiac; Random Allocation; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Time Factors; Ventricular Function, Left

The purpose of this study was to test for the presence of liver hypoxia and recovery after reperfusion when blood alcohol levels (BAL) are high. Male rats were fed ethanol intragastrically at a constant rate for 1 month. The pO(2) levels were then measured on the liver surface of these rats, in vivo during laparatomy under isoflurane anesthesia. To measure the response to acute hypoxia, the hepatic blood flow was clamped off at the porta hepatis. When the clamp was released, recovery from hypoxia was measured. A number of hypoxic-inducible genes in the liver were analyzed by means of quantitative RT-PCR as a measure of increased activation of hypoxia initiated transcription. The mRNA levels of genes for adrenomedullin, adrenergic receptor alpha, 1a and 1d, CDK inhibitor 1a, and erythropoietin were all significantly higher at the peaks than troughs. Expression of these same genes in the livers of control rats fed dextrose was lower than at the troughs. Although the mRNA level of the hypoxia-inducible factor (HIF-1alpha) was higher at the trough than at the peak, its protein concentration in the nuclear fraction was not increased at the troughs compared with the peaks. In fact, the nuclear protein level of HIF-1alpha at the peak was significantly higher than in control samples, which is consistent with the presence of hypoxia at the peaks. Further analysis of the HIF-alpha degradation regulation revealed that prolyl 4-hydroxylase (P4ha1) and von Hippel-Lindau syndrome homolog (Vhl) were both up-regulated at the troughs compared with the peaks. The liver surface oxygen levels at the peaks were reduced compared with the control samples. The pO(2) levels fell abruptly when the vessels at the porta hepatis were clamped. When the clamp was removed, allowing reperfusion of the liver, pO(2) returned to baseline levels in the control, and at the troughs but not at the peaks. These results support the hypothesis that hypoxia occurs at the peaks of the BAL cycle and recovery from ischemia is impaired at the peaks.

Topics: Administration, Oral; Adrenomedullin; Animals; Cell Cycle Proteins; Cyclin-Dependent Kinase Inhibitor p21; Disease Models, Animal; Erythropoietin; Ethanol; Hypoxia; Liver; Liver Diseases, Alcoholic; Male; Oligonucleotide Array Sequence Analysis; Oxygen; Peptides; Rats; Rats, Wistar; Receptors, Adrenergic; Reperfusion Injury; RNA, Messenger

Recent studies show that erythropoietin (EPO) plays a protective role in brain ischemia. In this condition, administration of EPO protects neurons from ischemic damage. Recently, it has been shown that in patients with chronic heart failure (CHF), EPO treatment improved cardiac function. In the present study we assessed the role of EPO and EPO-receptor (EPO-R) in the heart.. We studied the presence and functionality of the EPO-R in isolated rat hearts in the Langendorff set-up. Hearts were perfused for 20 min with 10 U/ml EPO or vehicle. Immunohistochemistry revealed the presence of the EPO-R on endothelial cells, fibroblasts and to a lesser extent cardiomyocytes. Furthermore, perfusion with EPO resulted in a 50% increase in the phosphorylated MAP kinases p42/p44. To evaluate the protective role of EPO in cardiac ischemia, we performed low-flow (0.6 ml/min) ischemia/reperfusion experiments in isolated rat hearts. Administration of EPO (10 U/ml) reduced the cellular damage by 56% (P<0.05) during reperfusion, diminished apoptosis by 15% (P<0.05) and resulted in a significantly improved recovery of left ventricular pressure (P=0.02) and coronary flow (P=0.01).. The present data suggest that a functional EPO-R is present in rat adult cardiac tissue and that exogenous EPO administration improves cardiac function after ischemia/reperfusion injury.

Topics: Animals; Coronary Circulation; Disease Models, Animal; Endothelial Cells; Erythropoietin; Fibroblasts; Humans; Immunohistochemistry; Mitogen-Activated Protein Kinases; Models, Cardiovascular; Phosphorylation; Rats; Receptors, Erythropoietin; Reperfusion Injury; Signal Transduction; Ventricular Function, Left

The aim of the study was investigate the effectiveness of recombinant human erythropoietin (r-Hu-EPO) in attenuating the severity of experimental liver ischemic injury in fetal rats.. The animals were divided randomly into four groups. In the control group, fetal whole liver tissue was taken immediately after laparotomy from pregnant animals. In the ischemia-reperfusion (I/R) group, tissue samples were taken immediately after I/R injury. In the vehicle group, 0.4 ml of human serum albumin solution and in the treatment group, r-Hu-EPO (5000 IU/kg) in 0.4 ml of human serum albumin solution were given intraperitoneally, 30 min before I/R injury, as a single dose. Thiobarbituric acid-reactive substances (TBARS) were estimated to demonstrate lipid peroxidation.. Lipid peroxidation byproducts increased after I/R injury. Administration of r-Hu-EPO reduced TBARS after I/R injury.. Further investigations are needed to understand the mechanism of the hepatoprotective effect of erythropoietin and the clinical importance of ischemic liver injury in the fetus.

Topics: Animals; Drug Administration Schedule; Erythropoietin; Female; Fetus; Humans; Injections, Intraperitoneal; Lipid Peroxidation; Liver; Pregnancy; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury; Thiobarbituric Acid Reactive Substances

Topics: Animals; Erythropoietin; Humans; Kidney; Rats; Reperfusion Injury

The aim of this study was to show the effect of erythropoietin on ischemia-reperfusion-induced oxidative damage in fetal rat brain.. Fetal brain ischemia was induced by clamping the utero-ovarian artery bilaterally for 20 min, and reperfusion was achieved by removing the clamps for 30 min. The control group was made up of non-injured rats that were 19 days pregnant. In the ischemia-reperfusion group no treatment was given, while 0.4 ml of human serum albumin solution and 5,000 U/kg recombinant human erythropoietin (r-Hu-EPO) were administered in the vehicle and treatment groups 30 min before ischemia-reperfusion injury. Lipid peroxidation in the brain tissue was determined as the concentration of thiobarbituric acid-reactive substances (TBARS) for each fetal rat. A one-way analysis of variance and the post-hoc test were used for statistical analysis.. TBARS increased to statistically significantly higher levels in fetal rat brain after ischemia-reperfusion injury than were found in the control group. Recombinant human erythropoietin prevented the increase in TBARS after ischemia-reperfusion injury.. Recombinant human erythropoietin has been shown to have neuroprotective effect in intrauterine ischemia-reperfusion-induced fetal brain damage in rats.

Topics: Animals; Brain; Erythropoietin; Female; Humans; Lipid Peroxidation; Oxidation-Reduction; Random Allocation; Rats; Rats, Wistar; Recombinant Proteins; Reperfusion Injury

Preconditioning phenomena provide evidence for adaptive responses to ischemia that have important implications for treatment/prevention of myocardial infarction. Hypoxia-inducible factor 1 (HIF-1) mediates adaptive transcriptional responses to hypoxia/ischemia.. Exposure of wild-type mice to intermittent hypoxia resulted in protection of isolated hearts against ischemia-reperfusion injury 24 hours later. Cardiac protection induced by intermittent hypoxia was lost in Hif1a+/- mice heterozygous for a knockout allele at the locus encoding HIF-1alpha. Erythropoietin (EPO) mRNA expression was induced in kidneys of wild-type mice subjected to intermittent hypoxia, resulting in increased plasma EPO levels. EPO mRNA expression was not induced in Hif1a+/- mice. EPO administration to rats increased functional recovery and decreased apoptosis in isolated hearts subjected to ischemia-reperfusion 24 hours later.. Hearts isolated from rodents subjected to intermittent hypoxia or EPO administration are protected against postischemic injury. Cardiac protection induced by intermittent hypoxia is critically dependent on Hif1a gene dosage. Our data suggest that additional studies to evaluate therapeutic applications of EPO administration are warranted.

Topics: Animals; Erythropoietin; Gene Expression; Heart; Heterozygote; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; In Vitro Techniques; Ischemic Preconditioning, Myocardial; Kidney; Male; Mice; Mice, Mutant Strains; Myocardium; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Rats; Rats, Sprague-Dawley; Receptors, Erythropoietin; Reperfusion Injury; RNA, Messenger; Time Factors; Transcription Factors

Improving the ability of the kidney to tolerate ischemic injury has important implications. We investigated the effect of recombinant human erythropoietin (rHuEPO) treatment on subsequent ischemia/reperfusion (I/R) injury and evaluated the role of heat shock protein (HSP) 70 in rHuEPO-induced renal protection. rHuEPO (3000 U/kg) was administered 24 h before I/R injury, and rats were killed at 24, 48, and 72 h after I/R injury. Pretreatment of rHuEPO resulted in the following: i) decreased serum creatinine level; ii) decreased tubular cell apoptosis and necrosis, measured by DNA fragmentation analysis and TUNEL staining and histomorphological criteria; iii) decreased tubular cell proliferation as determined by proliferating cell nuclear antigen expression; iv) increased bcl-2 protein and decreased caspase 3 activity; and v) decreased JNK expression. rHuEPO treatment increased HSP70 expression in a dose-dependent manner in normal rat kidneys, and inhibition of HSP70 expression by quercetin eliminated the renoprotective effect of rHuEPO in ischemic kidneys. Our study demonstrates that rHuEPO has a protective effect on subsequent I/R injury and that this effect is associated with induction of HSP70. Our study provides a new avenue for therapy to prevent renal damage after I/R injury.

Topics: Animals; Apoptosis; Erythropoietin; HSP70 Heat-Shock Proteins; Ischemic Preconditioning; Kidney; Kidney Diseases; Mitogen-Activated Protein Kinases; Models, Biological; Rats; Recombinant Proteins; Reperfusion Injury; Signal Transduction

Erythropoietin (EPO) plays an important role in the brain's response to neuronal injury. Systemic administration of recombinant human EPO (rhEPO) protects neurons from injury after middle cerebral artery occlusion, traumatic brain injury, neuroinflammation, and excitotoxicity. Protection is in part mediated by antiapoptotic mechanisms. We conducted parallel studies of rhEPO in a model of transient global retinal ischemia induced by raising intraocular pressure, which is a clinically relevant model for retinal diseases. We observed abundant expression of EPO receptor (EPO-R) throughout the ischemic retina. Neutralization of endogenous EPO with soluble EPO-R exacerbated ischemic injury, which supports a crucial role for an endogenous EPO/EPO-R system in the survival and recovery of neurons after an ischemic insult. Systemic administration of rhEPO before or immediately after retinal ischemia not only reduced histopathological damage but also promoted functional recovery as assessed by electroretinography. Exogenous EPO also significantly diminished terminal deoxynucleotidyltransferase-mediated dUTP end labeling labeling of neurons in the ischemic retina, implying an antiapoptotic mechanism of action. These results further establish EPO as a neuroprotective agent in acute neuronal ischemic injury.

Topics: Animals; Erythropoietin; Eye; In Situ Nick-End Labeling; Male; Neurons; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Receptors, Erythropoietin; Reperfusion Injury; Retina; Solubility

The anemia associated with acute renal failure (ARF) is currently treated with blood transfusions, while the anemia of chronic renal failure is treated with recombinant erythropoietin (EPO). We hypothesized that EPO treatment during ARF could rapidly improve hemoglobin levels and be a useful therapeutic approach. In addition, as tubular epithelial cells have EPO receptors that can mediate proliferation, enhanced recovery of renal function may occur with EPO use.. An established rat model of ischemic ARF was studied, using either moderate or severe ischemia. EPO was administered in a dose of 500 or 3000 U/kg starting at time of ischemia. Hematocrit (Hct), serum creatinine, reticulocyte count, and mortality rate were measured.. EPO treatment led to a rapid and significant increase in Hct at 48 and 72 hours after moderate ischemic renal reperfusion injury (IRI) in EPO (500 U/kg)-treated rats compared with control (saline treated) rats (mean +/- SE; 45.6 +/- 0.3% vs. 42.0 +/- 1.0%, P < 0.01) and (46.6 +/- 0.3 vs. 41.0 +/- 1.0, P < 0.01, N = 3 per group). In severe renal IRI, EPO treatment also led to significantly increased Hct at 48 (40.0 +/- 4.4% vs. 36.8 +/- 0.3%, P < 0.01, N = 3 per group) and 72 hours (43.5 +/- 1.5% vs. 34.7 +/- 2.3%, P < 0.01, N = 3 per group). Higher dose (3000 U/kg) EPO led to a more pronounced Hct increase after severe IRI at 48 hours compared with the 500 U/kg dose (43.5 +/- 0.3 vs. 40.3 +/- 0.3, P < 0.01, N = 3 per group). EPO treatment during moderate or severe renal IRI did not change the course of the renal dysfunction. EPO treatment (N = 19) had a significant protective effect on mortality during severe IRI. In addition, loss of body weight during ARF was not affected by EPO therapy.. Recombinant EPO can rapidly increase Hct and improve mortality during ARF. Human studies are warranted to evaluate the clinical applicability of this important finding.

Topics: Acute Kidney Injury; Anemia; Animals; Body Weight; Creatinine; Erythropoietin; Ischemia; Male; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Renal Circulation; Reperfusion Injury; Reticulocyte Count; Severity of Illness Index