Transcriptomic and bioinformatics analysis of the mechanism by which erythropoietin promotes recovery from traumatic brain injury in mice

Recent studies have found that erythropoietin promotes the recovery of neurological function after traumatic brain injury.However,the precise mechanism of action remains unclea r.In this study,we induced moderate traumatic brain injury in mice by intrape ritoneal injection of erythro poietin for 3 consecutive days.RNA sequencing detected a total of 4065 differentially expressed RNAs,including 1059 mRNAs,92 microRNAs,799 long non-coding RNAs,and 2115circular RNAs.Kyoto Encyclopedia of Genes and Genomes and Gene Ontology analyses revealed that the coding and non-coding RNAs that were differentially expressed after traumatic brain injury and treatment with erythropoietin play roles in the axon guidance pathway,Wnt pathway,and MAPK pathway.Constructing competing endogenous RNA networks showed that regulatory relationship between the differentially expressed non-coding RNAs and mRNAs.Because the axon guidance pathway was repeatedly enriched,the expression of Wnt5a and Ephb6,key factors in the axonal guidance pathway,was assessed.Ephb6 expression decreased and Wnt5a expression increased after traumatic brain injury,and these effects were reversed by treatment with erythro poietin.These findings suggest that erythro poietin can promote recove ry of nerve function after traumatic brain injury through the axon guidance pathway.

Evaluation of Hemoglobin and Serum Erythropoietin Levels in Patients with Polycythemia Vera and Secondary Polycythemia

Objective: This study compares hemoglobin and erythropoietin levels in patients with polycythemia vera and secondary polycythemia. Study Design: A retrospective cross-sectional study evaluating the serum erythropoietin and hemoglobin levels in patients with polycythemia vera and secondary polycythemia. This study was performed simultaneously in Texas state of the U.S. and Fars Province in Iran. Methods: Hemoglobin, hematocrit and erythropoietin test results were collected from patients aged 19 to 75 years who were diagnosed with polycythemia vera and secondary polycythemia. Patients records with history of thrombocythemia, congestive heart failure, dyspnea, anemia and pregnant woman were excluded from study. Patients in each decade of life were examined in separate groups, so that changes in hemoglobin related to aging did not affect the research results. Results: 75% of the patients were men, and 25% were women. A total of 1580 patients were analyzed in this study. 57.3% of patients in UTMB and 38.8 patients in Iran have hemoglobin level above 17 mg/dl. 74% of patients in UTMB and 88% of patients in Iran have erythropoietin below 10 IU/mL. Polycythemia in UTMB was more common in people over 50 and in Iran in patients under 50 years old. The serum hemoglobin and erythropoietin levels in patients with polycythemia vera were not significantly different in compare to secondary polycythemia patients. Data showed that there were 84 polycythemia patients per 100,000 people. The results of this study in UTMB and Iran showed that 4.5% and 7%, respectively, of patients with polycythemia had a positive JAK2 test. Conclusion: Low erythropoietin levels may not be helpful in differentiating polycythemia vera from secondary polycythemia. .

Desferoxamine preconditioning protects against cerebral ischemia in rats by inducing expressions of hypoxia inducible factor 1α and erythropoietin

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

红细胞生成素简介(Erythropoietin.EPO)

红细胞生成素是机体生理条件下以及失血后调控红细胞生成的主要生长刺激因子,在机体红细胞生成中具有重要作用。本文对近年来红细胞生成素在来源、分子结构、信号传导机制、生物学作用及临床应用等方面的研究进行了简单概述。

Protective effect of erythropoietin against 1-methyl-4-phenylpyridinium-induced neurodegenaration in PC12 cells

Objective The neuroprotective effect of erythropoietin （EPO） against 1-methyl-4-phenylpyridinium （MPP^＋）- induced oxidative stress in cultured PC12 cells, as well as the underlying mechanism, were investigated. Methods PC12 ceils impaired by MPP^＋ were used as the cell model of Parkinson＇s disease. Methyl thiazolyl tetrazolium （MTT） was used to assay the viability of the PC12 cells exposed to gradient concentrations of EPO, and the terminal deoxynucleotidyl transferase dUTP nick end labelling （TUNEL） assay was used to analyze the apoptosis ratio of PC 12 cells. The expression of Bcl-2 and Bax in PC 12 cells were examined by Western blot, and the reactive oxygen species （ROS）, the mitochondrial transmembrane potential and the activity of caspase-3 in each group were detected by spectrofluorometer. Results Treatment of PC12 cells with MPP^＋ caused the loss of cell viability, which may be associated with the elevation in apoptotic rate, the formation of ROS and the disruption of mitochondrial transmembrane potential. It was also shown that MPP＋ significantly induced the upregulation of Bax/Bcl-2 ratio and the activation of caspase-3. In contrast, EPO significantly reversed these responses and had the maximum protective effect at 1 U/mL. Conclusion The inhibitive effect of EPO on the MPP^＋ -induced cytotoxicity may be ascribed to its anti-oxidative property and anti-apoptotic activity, and EPO may provide a useful therapeutic strategy for treatment of neurodegenerative diseases such as Parkinson＇s disease.

促红细胞生成素干预牙髓损伤后修复性牙本质形成及骨形态发生蛋白2的表达

背景:促红细胞生成素具有抗炎、抗凋亡、促进骨缺损修复等作用,目前关于促红细胞生成素对牙髓损伤后修复性牙本质形成及其相关分子机制的研究较少。目的:探究促红细胞生成素对牙髓损伤后修复性牙本质形成的影响。方法:(1)动物实验:采用随机数字表法将32只大鼠随机分为对照组(n=16)与实验组(n=16),实验组牙髓损伤处使用含促红细胞生成素的胶原蛋白海绵直接盖髓,对照组露牙髓损伤处使用含PBS的胶原蛋白海绵直接盖髓,用玻璃离子粘固剂封闭窝洞;治疗2,4周后取上颌骨,采用免疫组化染色检测巢蛋白在牙本质中的表达,苏木精-伊红染色观察修复性牙本质生成。取4只SD大鼠上颌骨,免疫组化染色检测磨牙与切牙中促红细胞生成素表达。(2)细胞实验:从人牙髓组织、牙周韧带组织及牙龈组织中分别分离获取人牙髓细胞、人牙周韧带细胞和人牙龈成纤维细胞,采用RT-PCR检测促红细胞生成素mRNA表达。在成牙本质细胞诱导或未诱导分化条件下,采用RT-PCR检测人牙髓细胞中促红细胞生成素、牙本质涎磷蛋白、牙本质基质蛋白1、巢蛋白m RNA表达。在成牙本质细胞诱导或未诱导分化条件下,下调促红细胞生成素表达或外源性给予促红细胞生成素干预后,采用RT-PCR检测人牙髓细胞牙本质涎磷蛋白、牙本质基质蛋白1 mRNA相对表达,茜素红S染色检测矿化结节形成,RT-PCR与Western blotting检测骨形态发生蛋白2 mRNA与蛋白表达。结果与结论:(1)动物实验:与对照组相比,实验组治疗2,4周后的修复性牙本质生成量更大,牙本质中巢蛋白表达更高。促红细胞生成素在大鼠上颌第一磨牙牙髓、成牙本质细胞层和牙周膜中呈弱阳性表达,在成牙本质细胞中呈强阳性表达。(2)细胞实验:与其他两种细胞相比,人牙髓细胞中的促红细胞生成素m RNA表达更高。与未诱导分化相比,成牙本质细胞诱导分化下人牙髓细胞中牙本质涎磷蛋白、牙本质基质蛋白1、巢蛋白、促红细胞生成素与骨形态发生蛋白2 mRNA表达升高,矿化结节生成增加。在成牙本质细胞诱导分化下,下调促红细胞生成素表达后人牙髓细胞中牙本质涎磷蛋白、牙本质基质蛋白1、巢蛋白、骨形态发生蛋白2 mRNA表达与矿化结节生成均降低,骨形态发生蛋白2蛋白表达降低;外源性给予促红细胞生成素干预后,人牙髓细胞中上述指标表达均升高。(3)结果表明:促红细胞生成素对牙本质形成具有一定的促进作用。

Neuroprotective effects of erythropoietin on neurodegenerative and ischemic brain diseases: the role of erythropoietin receptor

Erythropoietin （Epo） is a fundamental hormone in the regulation of hematopoiesis, and other secondary roles mediated by the binding of the hormone to its specific receptor （EpoR）, which leads to an activation of key signaling pathways that induce an increase in cell differentiation, apoptosis control and neuroprotection. It has been suggested that their ftmction depends on final conformation of glycosylations, related with affinity to the receptor and its half-life. The presence of EpoR has been reported in different tissues including central nervous system, where it has been demonstrated to exert a neuroprotective function against oxidative stress conditions, such as ischemic injury and neurodegenerative diseases. There is also evidence of an increase in EpoR expression in brain cell lysates of Alzheimer＇s patients with respect to healthy patients. These results are related with extensive in vitro experimental data of neuroprotection obtained from cell lines, primary cell cultures and hippocampal slices. Additionally, this data is correlated with in vivo experiments （water maze test） in mouse models of Alzheimer＇s disease where Epo treatment improved cognitive function. These stud- ies support the idea that receptor activation induces a neuroprotective effect in neurodegenerative disorders including dementias, and especially Alzheimer＇s disease. Taken together, available evidence suggests that Epo appears to be a central element for EpoR activation and neuroprotective properties in the central nervous system. In this review, we will describe the mechanisms associated with neuroprotection and its relation with the activation of EpoR in order with identify new targets to develop pharmacological strategies.

Erythropoietin ameliorates early ischemia-reperfusion injury following the Pringle maneuver

AIM: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. METHODS: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. RESULTS: There were no increases in Hb and Ht in the EPO1 and EPO2 groups. AST was signif icantly 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 signif icantly lower in EPO1 than in STRD and EPO2 on Day 0 (P=0.020) and Day 1 (P=0.004). There were no signif icant inter-group differences in the levels of LDH and lactate. IL-6 was signif icantly lower in EPO1 than in STRD and EPO2 on Day 0 (P=0.0036) and Day 1 (P=0.0451). TNF-α was signif icantly lower in EPO1 than in STRD and EPO2 on Day 0 (P=0.0006) and Day 1 (P<0.0001). Furthermore, hospitalization was signif icantly shorter in EPO1 and EPO2 than in STRD.CONCLUSION:Epo has greater potential than steroids to ameliorate IR/I after the PM. Epo at a dose of 30000 U, administered before PM and just after surgery, yields better results.

Neuroprotection of Erythropoietin and Methylprednisolone against Spinal Cord Ischemia-Reperfusion Injury

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.

Neurotherapeutic potential of erythropoietin after ischemic injury of the central nervous system

Erythropoietin (EPO) is one of the most successful biopharmaceuticals in history and is used for treating anemia of different origins. However, it became clear that EPO could also work in a neuroprotective, antiapoptotic, antioxidative, angiogenetic and neurotropic way. It causes stimulation of cells to delay cell apoptosis, especially in the central nervous system. In rodent models of focal cerebral ischemia, EPO showed an impressive reduction of infarct size by 30% and improvement of neurobehavioral outcome by nearly 40%. A large animal model dealing with ischemia and reperfusion of the spinal cord showed that EPO could reduce the risk of spinal cord injury significantly. In addition, some clinical studies tested whether EPO works in real live clinical settings. One of the most promising studies showed the innocuousness and improvements in follow-up, outcome scales and in infarct size, of EPO-use in humans suffering from ischemic stroke. Another study ended unfortunately in a negative outcome and an increased overall death rate in the EPO group. The most possible reason was the involvement of patients undergoing simultaneously systemic thrombolysis with recombinant tissue plasminogen activator. An experimental study on rats demonstrated that administration of EPO might exacerbate tissue plasminogen activator-induced brain hemorrhage without reducing the ischemic brain damage. This case shows clearly how useful animal models can be to check negative side effects of a treatment before going into clinical trials. Other groups looked in human trials at the effects of EPO on the outcome after ischemic stroke, relation to circulating endothelial progerdtor cells, aneurysmal subarachnoid hemorrhage, traumatic brain injury, hemoglobin transfusion thresholds and elective first-time coronary artery bypass surgery. Most of the results were pos-让ive, but are based mostly on small group sizes. However, some of the most neglected facts when focusing on experimental setups of ischemia of the central nervous system are issues like age and comorbidities. It might be extremely worthy to consider these points for future projects, because EPO might influence all these factors.

The Erythropoietin Receptor: Dimerization and Intracellular Signal Transduction

Erythropoietin (EPO), a 34 kD glycopro-tein, is the principal growth factor regulating theproduction of circulating erythrocytes; EPO isessential for committed CFU - E erythroid pro-genitors to divide several times and then to dif-ferentiate into erythrocytes. Like most receptorsfor hematopoietic growth factors, the erythro-poietin receptor (EPO - R) is a type I trans-membrane protein and a member of the cytokinereceptor superfamily. These receptors containfour conserved cysteines and a Trp - Ser - X -

Pretreatment with erythropoietin reduces hepatic ischemia-reperfusion injury

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSION: These results demonstrate that EPO can up-regulate HO-1 in liver tissues and accordingly decrease hepatic injury through its anti-inflammatory property. (Hepatobiliary Pancreat Dis Int 2009; 8: 294-299)

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

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

Effects of erythropoietin on the expression of tumor necrosis factor-alpha and Bax after facial nerve axotomy in rats

This study sought to evaluate the effect of high-dose erythropoietin （EPO; 5 000 IU/kg） on the expression of tumor necrosis factor-alpha （TNF-α） and Bax in the facial nucleus after facial nerve transection in rats. A total of 42 Wistar rats of both genders were used in this study, and 40 rats were randomly divided into 2 groups： EPO group and model group. The EPO group was treated with EPO once a day for 5 days at a dose of 5 000 IU/kg body weight. The model group was treated with saline of the same amount. At day 3 after EPO （or saline） treatment, the right facial nerves of the 40 rats were transected at the level of the stylomastoid foramen, with the left sides untreated. The remaining 2 rats that did not undergo axotomy served as the control group. The surviving motor neurons in operated rats were counted in coronal paraffin sections of the facial nucleus. The expression of TNF-a and Bax in the facial nucleus was detected by immunohistochemical staining at days 3, 7, 14, 21, and 28 after axotomy. At days 14, 21, and 28 after facial nerve axotomy, a significantly greater proportion of facial motor neurons survived in the EPO group than in the model group. After axotomy, the expression of TNF-a and Bax increased in motor neurons in both the EPO and the model groups. TNF-o expression reached its peak level at day 14 after axotomy, while Bax expression reached its peak level at day 21. TNF-α expression was much lower in the EPO group than in the model group at all time points. No significant difference in Bax expression was found between the EPO and the model groups. These results indicate that high-dose EPO treatment attenuates the increase in TNF-α expression in the facial nucleus and reduces the loss of motor neurons after facial nerve transection in rats. However, high-dose EPO treatment has little effect on Bax expression.

Circulating autoantibodies to endogenous erythropoietin are associated with chronic hepatitis C virus infection-related anemia

BACKGROUND： Chronic hepatitis C virus （HCV） infection is associated with autoimmune phenomena and is often complicated by anemia. Circulating autoantibodies to endogenous erythropoietin （anti-EPO） have been detected in patients with chronic viral infections and were correlated to anemia. The present study aimed to determine anti-EPO prevalence in pa- tients with chronic HCV infection and investigate its possible association with anemia. METHODS： Ninety-three consecutive patients （62 males and 31 females） with chronic HCV infection, who had never re- ceived antiviral therapy or recombinant EPO, were enrolled in the study. Circulating anti-EPO were detected in the serum by using an ELISA assay. Quantitative determination of serum EPO levels was done by radioimmunoassay. HCV RNA viral load t and genotype sequencing were also performed. RESULTS： Circulating anti-EPO were detected in 10.8% of HCV-infected patients and the prevalence of anti-EPO was significantly higher in patients with anemia （19.4% vs 5.3%, P=0.040） compared to that in those without anemia. Compared to anti-EPO negative cases, anti-EPO positive patients had higher frequency of anemia （70.0% vs 34.9%, P=0.030）, lower EPO concentrations （median 16.35 vs 30.65 mU/mL, P=0.005）, and higher HCV RNA viral load （median 891.5x103 vs 367.5x 103 IU/mL, P=0.016）. In multivariate regression anal- ysis the presence of anti-EPO remained an independent predictor of anemia （adjusted OR： 14.303, 95% CI： 1.417-36.580, P=0.024）. EPO response to anemia was less prominent among anti-EPO positive patients （P=0.001）. CONCLUSIONS： Circulating anti-EPO are detected in a significant proportion of treatment-naive HCV-infected patients and are independently associated with anemia, suggesting a further implication of autoimmunity in the pathophysiology of HCV-related anemia.

Erythropoietin gene transfer into rat testes by in vivo electropo-ration may reduce the risk of germ cell loss caused by cryptorchidism

Aim： To investigate the effects of rat Erythropoietin （Epo） on spermatogenesis by transferring rat Epo gene into cryptorchid testes by means of in vivo electroporation. Methods： Sprague-Dawley rats with surgically-induced unilateral cryptorchidism were divided into three groups： the first group was given intratesticular injections of pCAGGS-Epo （pCAGGS-Epo group）, the second group was given intratesticular injections of pCAGGS （pCAGGS group）, and the third group were given intratesticular injections of phosphate-buffered saline （PBS group）. At the same time, square electric pulses of 30 V were applied six times with a time constant of 100 ms. One or two weeks after injection, each testis was weighed and the ratio of the total number of germ cells to that of Sertoli cells （G/S ratio） was calculated to evaluate the impairment of spermatogenesis. Ten testes taken from each of the three groups were examined at each time point. Results： The testicular weight after the injection of pCAGGS-Epo or pCAGGS control plasmid was （0.85 ± 0.08） g and （0.83 ± 0.03） g, respectively, at week 1 （P = 0.788） and （0.62 ± 0.06） g and （0.52 ± 0.02） g, respectively, at week 2 （P = 0.047）. At week 1, spermatids and sperm were more abundant in testes with pCAGGS-Epo than those in the control testes. At week 2, spermatids and sperm were hardly detected in either group. The G/S ratio was 23.27 ± 6.80 vs. 18.63 ± 5.30 at week 1 （P = 0.0078） and 7.16 ± 3.06 vs. 6.05 ± 1.58 at week 2 （P = 0.1471）, respectively. Conclusion： The transfer of Epo to rat testes by in vivo electroporation may reduce the risk of the germ cell loss caused by cryptorchidism.

Erythropoietin reduces apoptosis of brain tissue cells in rats after cerebral ischemia/reperfusion injury:a characteristic analysis using magnetic resonance imaging

Some in vitro experiments have shown that erythropoietin （EPO） increases resistance to apoptosis and facilitates neuronal survival follow- ing cerebral ischemia. However, results from in vivo studies are rarely reported. Perfusion-weighted imaging （PWI） and diffusion-weighted imaging （DWI） have been applied successfully to distinguish acute cerebral ischemic necrosis and penumbra in living animals; therefore, we hypothesized that PWI and DWI could be used to provide imaging evidence in vivo for the conclusion that EPO could reduce apoptosis in brain areas injured by cerebral ischemia/reperfusion. To validate this hypothesis, we established a rat model of focal cerebral ischemia/ reperfusion injury, and treated with intra-cerebroventricular injection of EPO （5,000 U/kg） 20 minutes before injury. Brain tissue in the ischemic injury zone was sampled using MRI-guided localization. The relative area of abnormal tissue, changes in PWI and DWI in the ischemic injury zone, and the number of apoptotic cells based on TdT-mediated dUTP-biotin nick end-labeling （TUNEL） were assessed. Our findings demonstrate that EPO reduces the relative area of abnormally high signal in PWI and DWI, increases cerebral blood volume, and decreases the number of apoptotic cells positive for TUNEL in the area injured by cerebral ischemia/reperfusion. The experiment pro- vides imaging evidence in vivo for EPO treating cerebral ischemia/reperfusion injury.

Erythropoietin therapy after out-of-hospital cardiac arrest:A systematic review and meta-analysis

AIM To assess safety and efficacy of early erythropoietin(Epo) administration in patients with out-of-hospital cardiac arrest(OHCA).METHODS A systematic literature search was performed using PubM ed,MEDLINE,EMBASE,EBSCO,CINAHL,Web of Science and Cochrane databases,of all studies published from the inception through October 10,2016.Inclusion criteria included:(1) Adult humans with OHCA and successful sustained return of spontaneous circulation;and(2) studies including mortality/brain death,acute thrombotic events as their end points.Primary efficacyoutcome was "brain death or Cerebral Performance Category(CPC) score of 5".Secondary outcomes were "CPC score 1,and 2-4","overall thrombotic events" and "acute coronary stent thrombosis".RESULTS We analyzed a total of 606 participants(n = 276 received Epo and n = 330 with standard of care alone) who experienced OHCA enrolled in 3 clinical trials.No significant difference was observed between the Epo and no Epo group in brain death or CPC score 5(OR = 0.77;95%CI:0.42-1.39),CPC score 1(OR = 1.16,95%CI:0.82-1.64),and CPC score 2-4(OR = 0.77,95%CI:0.44-1.36).Epo group was associated with increased thrombotic complications(OR = 2.41,95%CI:1.26-4.62) and acute coronary stent thrombosis(OR = 8.16,95%CI:1.39-47.99).No publication bias was observed.CONCLUSION Our study demonstrates no improvement in neurological outcomes and increased incidence of thrombotic events and acute coronary stent thrombosis in OHCA patients who were treated with Epo in addition to standard therapy.

Association of erythropoietin gene rs576236 polymorphism and risk of adrenal tumors in a Chinese population

Erythropoietin（EPO） is a circulating glycosylated protein hormone and has been implicated in the development and progression of non-hematopoietic tissue tumors.The objective of the present study was to determine if the EPO rs576236 polymorphism was associated with the risk of adrenal tumors.We genotyped the EPO rs576236 polymorphism in a case-control study of 288 adrenal tumor patients and 456 cancer-free controls by using the TaqMan method,and assessed the association between the polymorphism and the adrenal tumor risk by logistic regression.Furthermore,95%confidence interval（CI） was used to assess the genetic association between the polymorphism and the risk of adrenal tumor.Compared with the TT genotype,the TC genotype had a significantly increased risk of adrenal tumor[adjusted odds ratio（OR） = 1.24,95%CI = 1.12-2.22].Furthermore a significantly increased risk of adrenal tumor was found in the combined variant genotypes TC＋CC compared with the TT genotype（adjusted OR = 1.17,95%CI = 1.12-2.21）.Our present study suggests that the rs576236 polymorphism of EPO confers susceptibility to adrenal tumor in the Chinese population.

Cerebral Ischemic Tolerance Induced by 3-nitropropionic Acid Is Associated with Increased Expression of Erythropoietin in Rats

Summary： 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.