Advertisement

Intensive Care Medicine

, 37:1525 | Cite as

Comparison of carbamylated erythropoietin-FC fusion protein and recombinant human erythropoietin during porcine aortic balloon occlusion-induced spinal cord ischemia/reperfusion injury

  • Florian Simon
  • Angelika Scheuerle
  • Michael Gröger
  • Brigitta Vcelar
  • Oscar McCook
  • Peter Möller
  • Michael Georgieff
  • Enrico Calzia
  • Peter Radermacher
  • Hubert Schelzig
Experimental

Abstract

Purpose

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.

Methods

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.

Results

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.

Conclusions

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.

Keywords

Motor evoked potentials Lower limb reflexes Nissl staining TUNEL staining Cytokines 8-Isoprostanes 

Notes

Acknowledgment

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.

References

  1. 1.
    Gelman S (1995) The pathophysiology of aortic cross-clamping and unclamping. Anesthesiology 82:1026–1060PubMedCrossRefGoogle Scholar
  2. 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
  3. 3.
    Sönmez A, Kabakçi B, Vardar E, Gürel D, Sönmez U, Orhan YT, Açikel U, Gökmen N (2007) Erythropoietin attenuates neuronal injury and potentiates the expression of pCREB in anterior horn after transient spinal cord ischemia in rats. Surg Neurol 68:297–303PubMedCrossRefGoogle Scholar
  4. 4.
    Simon F, Scheuerle A, Calzia E, Bassi G, Oter S, Duy CN, Kick J, Brückner UB, Radermacher P, Schelzig H (2008) Erythropoietin during porcine aortic balloon occlusion-induced ischemia/reperfusion injury. Crit Care Med 36:2143–2150PubMedCrossRefGoogle Scholar
  5. 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
  6. 6.
    Nagai T, Akizawa T, Kohjiro S, Koiwa F, Nabeshima K, Niikura K, Kino K, Kanamori N, Kinugasa E, Ideura T (1996) rHuEPO enhances the production of plasminogen activator inhibitor-1 in cultured endothelial cells. Kidney Int 50:102–107PubMedCrossRefGoogle Scholar
  7. 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. 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
  9. 9.
    Brines M, Cerami A (2008) Erythropoietin-mediated tissue protection: reducing collateral damage from the primary injury response. J Int Med 264:405–432CrossRefGoogle Scholar
  10. 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
  11. 11.
    Adembri C, Massagrande A, Tani A, Miranda M, Margheri M, De Gaudio R, Pellegrini-Giampietro DE (2008) Carbamylated erythropoietin is neuroprotective in an experimental model of traumatic brain injury. Crit Care Med 36:975–978PubMedCrossRefGoogle Scholar
  12. 12.
    Lapchak PA, Kirkeby A, Zivin JA, Sager TN (2008) Therapeutic window for nonerythropoietic carbamylated-erythropoietin to improve motor function following multiple infarct ischemic strokes in New Zealand white rabbits. Brain Res 1238:208–214PubMedCrossRefGoogle Scholar
  13. 13.
    Nijboer WN, Ottens PJ, van Dijk A, van Goor H, Ploeg RJ, Leuvenink HGD (2010) Donor pretreatment with carbamylated erythropoietin in a brain death model reduces inflammation more effectively than erythropoietin while preserving renal function. Crit Care Med 38:1155–1161PubMedCrossRefGoogle Scholar
  14. 14.
    Schriebl K, Trummer E, Lattenmayer C, Weik R, Kunert R, Müller D, Katinger H, Vorauer-Uhl K (2006) Biochemical characterization of rhEPO-Fc fusion protein expressed in CHO cells. Protein Expr Purif 49:265–275PubMedCrossRefGoogle Scholar
  15. 15.
    Simon F, Giudici R, Duy CN, Schelzig H, Oter S, Gröger M, Wachter U, Vogt J, Speit G, Szabó C, Radermacher P, Calzia E (2008) Hemodynamic and metabolic effects of hydrogen sulfide during porcine ischemia/reperfusion injury. Shock 30:359–364PubMedCrossRefGoogle Scholar
  16. 16.
    Kuriyama T, Latham LP, Horwitz LD, Reeves JT, Wagner WW Jr (1984) Role of collateral ventilation in ventilation-perfusion balance. J Appl Physiol 56:1500–1506PubMedGoogle Scholar
  17. 17.
    Nirmalan M, Willard TM, Edwards DJ, Little RA, Dark PM (2005) Estimation of errors in determining intrathoracic blood volume using the single transpulmonary thermal dilution technique in hypovolemic shock. Anesthesiology 103:805–812PubMedCrossRefGoogle Scholar
  18. 18.
    Papakostas JC, Matsagas MI, Toumpoulis IK, Malamou-Mitsi VD, Pappa LS, Gkrepi C, Anagnostopoulos CE, Kappas AM (2006) Evolution of spinal cord injury in a porcine model of prolonged aortic occlusion. J Surg Res 133:159–166PubMedCrossRefGoogle Scholar
  19. 19.
    Blaisdell FW, Cooley DA (1962) The mechanism of paraplegia after temporary thoracic aortic occlusion and its relationship to spinal fluid pressure. Surgery 51:351–355PubMedGoogle Scholar
  20. 20.
    Wadouh F, Lindemann EM, Arndt CF, Hetzer R, Borst HG (1984) The arteria radicularis magna anterior as a decisive factor influencing spinal cord damage during aortic occlusion. J Thorac Cardiovasc Surg 88:1–10PubMedGoogle Scholar
  21. 21.
    Weigang E, Luehr M, von Samson P, Hartert M, Goebel H, Wetzig M, Bernard V, Siegenthaler MP, Beyersdorf F (2005) Development of a special balloon occlusion device top prevent adverse events in high-risk patients during open aortic surgery. Eur Surg Res 37:204–209PubMedCrossRefGoogle Scholar
  22. 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
  23. 23.
    Meylaerts SA, De Haan P, Kalkman CJ, Jaspers J, Vanicky I, Jacobs MJ (2000) Prevention of paraplegia in pigs by selective segmental artery perfusion during aortic crossclamping. J Vasc Surg 32:160–170PubMedCrossRefGoogle Scholar
  24. 24.
    Lips J, de Haan P, Bouma GJ, Holman R, van Dongen E, Kalkman CJ (2005) Continuous monitoring of cerebrospinal fluid oxygen tension in relation to motor evoked potentials during spinal cord ischemia in pigs. Anesthesiology 102:340–345PubMedCrossRefGoogle Scholar
  25. 25.
    Hauser B, Kick J, Iványi Z, Asfar P, Ehrmann U, Muth CM, Albicini M, Wachter U, Vogt J, Bauer M, Brückner UB, Radermacher P, Bracht H (2006) Effects of 15-deoxy-Δ12, 14-prostaglandin-J2 during hyperdynamic porcine endotoxemia. Intensive Care Med 3:759–765CrossRefGoogle Scholar
  26. 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
  27. 27.
    Maier C, Scheuerle A, Hauser B, Schelzig H, Szabó C, Radermacher P, Kick J (2007) The selective PARP-1 inhibitor INO1001 reduces spinal cord injury during porcine aortic cross-clamping-induced ischemia/reperfusion injury. Intensive Care Med 33:845–850PubMedCrossRefGoogle Scholar
  28. 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
  29. 29.
    Kaptanoglu E, Solaroglu I, Okutan O, Surucu HS, Akbiyik F, Beskonakli E (2004) Erythropoietin exerts neuroprotection after acute spinal cord injury in rats: effect on lipid peroxidation and early ultrastructural findings. Neurosurg Rev 27:113–120PubMedCrossRefGoogle Scholar
  30. 30.
    King VR, Averill SA, Hewazy D, Priestley JV, Torup L, Michael-Titus AT (2007) Erythropoietin and carbamylated erythropoietin are neuroprotective following spinal cord hemisection in the rat. Eur J Neurosci 26:90–100PubMedCrossRefGoogle Scholar
  31. 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. 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
  33. 33.
    Lang-Lazdunski L, Heurteaux C, Mignon A, Mantz J, Widmann C, Desmonts J, Lazdunski M (2000) Ischemic spinal cord injury induced by aortic cross-clamping: prevention by riluzole. Eur J Cardiothorac Surg 18:174–181PubMedCrossRefGoogle Scholar
  34. 34.
    Wang Q, Ding Q, Zhou Y, Gou X, Hou L, Chen S, Zhu Z, Xiong L (2009) Ethyl pyruvate attenuates spinal cord ischemic injury with a wide therapeutic window through inhibiting high-mobility group box 1 release in rabbits. Anesthesiology 110:1279–1286PubMedCrossRefGoogle Scholar
  35. 35.
    Hu BR, Liu CL, Ouyang Y, Blomgren K, Siesjö BK (2000) Involvement of caspase-3 in cell death after hypoxia-ischemia declines during brain maturation. J Cereb Blood Flow Metab 20:1550–1556PubMedCrossRefGoogle Scholar
  36. 36.
    Kuluz JW, Prado R, He D, Zhao W, Dietrich WD, Watson B (2007) New pediatric model of ischemic stroke in infant piglets by photothrombosis: acute changes in cerebral blood flow, microvasculature, and early histopathology. Stroke 38:1932–1937PubMedCrossRefGoogle Scholar
  37. 37.
    Maiese K, Li F, Chong ZZ (2005) New avenues of exploration for erythropoietin. JAMA 293:90–95PubMedCrossRefGoogle Scholar
  38. 38.
    Moon C, Krawczyk M, Paik D, Coleman T, Brines M, Juhaszova M, Sollott SJ, Lakatta EG, Talan MI (2006) Erythropoietin, modified to not stimulate red blood cell production, retains its cardioprotective properties. J Pharmacol Exp Ther 316:999–1005PubMedCrossRefGoogle Scholar
  39. 39.
    Basu S, Eriksson M (1998) Oxidative injury and survival during endotoxemia. FEBS Lett 438:159–160PubMedCrossRefGoogle Scholar
  40. 40.
    Siems W, Quast S, Carluccio F, Wiswedel I, Hirsch D, Augustin W, Kraemer K, Hampl H, Sommerburg O (2003) Oxidative stress in cardio renal anemia syndrome: correlations and therapeutic possibilities. Clin Nephrol 60 Suppl 1:S22–S30PubMedGoogle Scholar
  41. 41.
    Nalos M, Asfar P, Ichai C, Radermacher P, Leverve XM, Fröba G (2003) Adenosinetriphosphate-magnesium chloride: relevance for intensive care. Intensive Care Med 29:10–18PubMedGoogle Scholar
  42. 42.
    Maharajh GS, Pascoe EA, Halliday WC, Grocott HP, Thiessen DB, Girling LG, Cheang MS, Mutch WA (1996) Neurological outcome in a porcine model of descending thoracic aortic surgery. Left atrial–femoral artery bypass versus clamp/repair. Stroke 27:2095–2101PubMedCrossRefGoogle Scholar

Copyright information

© Copyright jointly held by Springer and ESICM 2011

Authors and Affiliations

  • Florian Simon
    • 1
    • 2
  • Angelika Scheuerle
    • 3
  • Michael Gröger
    • 2
  • Brigitta Vcelar
    • 4
  • Oscar McCook
    • 2
  • Peter Möller
    • 3
  • Michael Georgieff
    • 2
  • Enrico Calzia
    • 2
  • Peter Radermacher
    • 2
  • Hubert Schelzig
    • 1
  1. 1.Abteilung für Thorax- und GefässchirurgieUniversitätsklinikumUlmGermany
  2. 2.Sektion Anästhesiologische Pathophysiologie und Verfahrensentwicklung, Klinik für AnästhesiologieUniversitätsklinikumUlmGermany
  3. 3.Abteilung PathologieUniversitätsklinikumUlmGermany
  4. 4.Polymun Scientific GmbHViennaAustria

Personalised recommendations