Comparison of carbamylated erythropoietin-FC fusion protein and recombinant human erythropoietin during porcine aortic balloon occlusion-induced spinal cord ischemia/reperfusion injury
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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.
KeywordsMotor evoked potentials Lower limb reflexes Nissl staining TUNEL staining Cytokines 8-Isoprostanes
This study was supported by the Deutsche Forschungsgemeinschaft (DFG Sche 899/2-3). Carbamylated erythropoietin fusion protein (cEPO-FC) was kindly provided by Polymun Scientific GmbH, Vienna, Austria. The authors are indebted to Andrea Söll and Tanja Schulz for skillful assistance.
Conflicts of interest
B. Vcelar is an employee responsible for preclinical research at Polymun Scientific GmbH (Vienna, Austria), a company involved in the commercial development of cEPO-FC, but holds no equity in that company nor related to the molecules investigated. The other authors declare that they have no competing interests at all.
- 2.Celik M, Gökmen N, Erbayraktar S, Akhisaroglu M, Konakc S, Ulukus C, Genc S, Genc K, Sagiroglu E, Cerami A, Brines M (2002) Erythropoietin prevents motor neuron apoptosis and neurologic disability in experimental spinal cord ischemic injury. Proc Natl Acad Sci USA 99:2258–2263PubMedCrossRefGoogle Scholar
- 5.Ehrenreich H, Weissenborn K, Prange H, Schneider D, Weimar C, Wartenberg K, Schellinger PD, Bohn M, Becker H, Wegrzyn M, Jähnig P, Herrmann M, Knauth M, Bähr M, Heide W, Wagner A, Schwab S, Reichmann H, Schwendemann G, Dengler R, Kastrup A, Bartels C, Stroke Trial Group EPO (2010) Recombinant human erythropoietin in the treatment of acute ischemic stroke. Stroke 40:647–656CrossRefGoogle Scholar
- 7.Brines M, Grasso G, Fiordaliso F, Sfacteria A, Ghezzi P, Fratelli M, Latini R, Xie QW, Smart J, Su-Rick CJ, Pobre E, Diaz D, Gomez D, Hand C, Coleman T, Cerami A (2004) Erythropoietin mediates tissue protection through an erythropoietin and common beta-subunit heteroceptor. Proc Natl Acad Sci USA 1:14907–14912CrossRefGoogle Scholar
- 8.Coleman TR, Westenfelder C, Tögel FE, Yang Y, Hu Z, Swenson L, Leuvenink HG, Ploeg RJ, d Uscio LV, Katusic ZS, Ghezzi P, Zanetti A, Kaushansky K, Fox NE, Cerami A, Brines M (2006) Cytoprotective doses of erythropoietin or carbamylated erythropoietin have markedly different procoagulant and vasoactive activities. Proc Natl Acad Sci USA 103:5965–5970PubMedCrossRefGoogle Scholar
- 10.Leist M, Ghezzi P, Grasso G, Bianchi R, Villa P, Fratelli M, Savino C, Bianchi M, Nielsen J, Gerwien J, Kallunki P, Larsen AK, Helboe L, Christensen S, Pedersen LO, Nielsen M, Torup L, Sager T, Sfacteria A, Erbayraktar S, Erbayraktar Z, Gokmen N, Yilmaz O, Cerami-Hand C, Xie QW, Coleman T, Cerami A, Brines M (2004) Derivatives of erythropoietin that are tissue protective but not erythropoietic. Science 305:239–242PubMedCrossRefGoogle Scholar
- 22.Meylaerts SA, De Haan P, Kalkman CJ, Lips J, De Mol BA, Jacobs MJ (1999) The influence of regional spinal cord hypothermia on transcranial myogenic motor-evoked potential monitoring and the efficacy of spinal cord ischemia detection. J Thorac Cardiovasc Surg 118:1038–1045PubMedCrossRefGoogle Scholar
- 26.Kick J, Hauser B, Bracht H, Albicini M, Oter S, Simon F, Ehrmann U, Garrel C, Sträter J, Brückner UB, Leverve XM, Schelzig H, Speit G, Radermacher P, Muth CM (2007) Effects of a cantaloupe melon extract/wheat gliadin biopolymer during aortic cross-clamping. Intensive Care Med 33:694–702PubMedCrossRefGoogle Scholar
- 28.Villa P, van Beek J, Larsen AK, Gerwien J, Christensen S, Cerami A, Brines M, Leist M, Ghezzi P, Torup L (2007) Reduced functional deficits, neuroinflammation, and secondary tissue damage after treatment of stroke by nonerythropoietic erythropoietin derivatives. J Cerebr Blood Flow Metab 27:552–563CrossRefGoogle Scholar
- 31.Simon F, Scheuerle A, Gröger M, Stahl B, Wachter U, Vogt J, Speit G, Hauser B, Möller P, Calzia E, Szabó C, Schelzig H, Georgieff M, Radermacher P, Wagner F (2011) Effects of intravenous sulfide during porcine aortic occlusion-induced kidney ischemia/reperfusion injury. Shock 35:156–163PubMedCrossRefGoogle Scholar
- 32.Kuluz J, Samdani A, Benglis D, Gonzalez-Brito M, Solano JP, Ramirez MA, Luqman A, De los Santos R, Hutchinson D, Nares M, Padgett K, He D, Huang T, Levi A, Betz R, Dietrich D (2010) Pediatric spinal cord injury in infant piglets: description of a new large animal model and review of the literature. J Spinal Cord Med 33:43–57PubMedGoogle Scholar