Molecular Medicine

, Volume 20, Issue 1, pp 516–526 | Cite as

Reduction of Cardiac Cell Death after Helium Postconditioning in Rats: Transcriptional Analysis of Cell Death and Survival Pathways

  • Gezina T. M. L. Oei
  • Michal Heger
  • Rowan F. van Golen
  • Lindy K. Alles
  • Moritz Flick
  • Allard C. van der Wal
  • Thomas M. van Gulik
  • Markus W. Hollmann
  • Benedikt Preckel
  • Nina C. Weber
Research Article


Helium, a noble gas, has been used safely in humans. In animal models of regional myocardial ischemia/reperfusion (I/R) it was shown that helium conditioning reduces infarct size. Currently, it is not known how helium exerts its cytoprotective effects and which cell death/survival pathways are affected. The objective of this study, therefore, was to investigate the cell protective effects of helium postconditioning by PCR array analysis of genes involved in necrosis, apoptosis and autophagy. Male rats were subjected to 25 min of ischemia and 5, 15 or 30 min of reperfusion. Semiquantitative histological analysis revealed that 15 min of helium postconditioning reduced the extent of I/R-induced cell damage. This effect was not observed after 5 and 30 min of helium postconditioning. Analysis of the differential expression of genes showed that 15 min of helium postconditioning mainly caused upregulation of genes involved in autophagy and inhibition of apoptosis versus I/R alone. The results suggest that the cytoprotective effects of helium inhalation may be caused by a switch from pro-cell-death signaling to activation of cell survival mechanisms, which appears to affect a wide range of pathways.

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  1. 1.
    Kramarow E, Lubitz J, Francis R. (2013) Trends in the coronary heart disease risk profile of middle-aged adults. Ann. Epidemiol. 23:31–4.CrossRefPubMedGoogle Scholar
  2. 2.
    Ovize M, et al. (2010) Postconditioning and protection from reperfusion injury: where do we stand? Position paper from the Working Group of Cellular Biology of the Heart of the European Society of Cardiology. Cardiovasc. Res. 87:406–23.CrossRefPubMedGoogle Scholar
  3. 3.
    Hamacher-Brady A, Brady NR, Gottlieb RA. (2006) The interplay between pro-death and pro-survival signaling pathways in myocardial ischemia/reperfusion injury: apoptosis meets autophagy. Cardiovasc. Drugs Ther. 20:445–62.CrossRefPubMedGoogle Scholar
  4. 4.
    Jain MV, et al. (2013) Interconnections between apoptotic, autophagic and necrotic pathways: implications for cancer therapy development. J. Cell. Mol. Med. 17:12–29.CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Konstantinidis K, Whelan RS, Kitsis RN. (2012) Mechanisms of cell death in heart disease. Arterioscler. Thromb. Vasc. Biol. 32:1552–62.CrossRefPubMedGoogle Scholar
  6. 6.
    Barach A. (1934) Use of helium as a new therapeutic gas. Proc. Soc. Exp. Bio. Med. 32:462–4.CrossRefGoogle Scholar
  7. 7.
    Moraa I, Sturman N, McGuire T, van Driel ML. (2013) Heliox for croup in children. Cochrane Database Syst. Rev. 12:CD006822.Google Scholar
  8. 8.
    Rodrigo G, Pollack C, Rodrigo C, Rowe BH. (2006) Heliox for nonintubated acute asthma patients. Cochrane Database Syst. Rev. 18:CD002884.Google Scholar
  9. 9.
    Bennett MH, Lehm JP, Mitchell SJ, Wasiak J. (2012) Recompression and adjunctive therapy for decompression illness. Cochrane Database Syst Rev. 5:CD005277.Google Scholar
  10. 10.
    Oei GT, Weber NC, Hollmann MW, Preckel B. (2010) Cellular effects of helium in different organs. Anesthesiology. 112:1503–10.CrossRefPubMedGoogle Scholar
  11. 11.
    Huhn R, et al. (2012) Age-related loss of cardiac preconditioning: impact of protein kinase A. Exp. Gerontol. 47:116–21.CrossRefPubMedGoogle Scholar
  12. 12.
    Pagel PS, Krolikowski JG. (2009) Transient metabolic alkalosis during early reperfusion abolishes helium preconditioning against myocardial infarction: restoration of cardioprotection by cyclosporin A in rabbits. Anesth. Analg. 108:1076–82.CrossRefPubMedGoogle Scholar
  13. 13.
    Pagel PS, Krolikowski JG, Amour J, Warltier DC, Weihrauch D. (2009) Morphine reduces the threshold of helium preconditioning against myocardial infarction: the role of opioid receptors in rabbits. J. Cardiothorac. Vasc. Anesth. 23:619–24.CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Pagel PS, et al. (2007) Noble gases without anesthetic properties protect myocardium against infarction by activating prosurvival signaling kinases and inhibiting mitochondrial permeability transition in vivo. Anesth. Analg. 105:562–9.CrossRefPubMedGoogle Scholar
  15. 15.
    Huhn R, et al. (2009) Helium-induced early preconditioning and postconditioning are abolished in obese Zucker rats in vivo. J. Pharmacol. Exp. Ther. 329:600–7.CrossRefPubMedGoogle Scholar
  16. 16.
    Heinen A, et al. (2008) Helium-induced preconditioning in young and old rat heart: impact of mitochondrial Ca(2+)-sensitive potassium channel activation. Anesthesiology. 109:830–6.CrossRefPubMedGoogle Scholar
  17. 17.
    Pagel PS, et al. (2008) Inhibition of glycogen synthase kinase or the apoptotic protein p53 lowers the threshold of helium cardioprotection in vivo: the role of mitochondrial permeability transition. Anesth. Analg. 107:769–75.CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Pagel PS, et al. (2008) The mechanism of helium-induced preconditioning: a direct role for nitric oxide in rabbits. Anesth. Analg. 107:762–8.CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Pagel PS, et al. (2008) Reactive oxygen species and mitochondrial adenosine triphosphate-regulated potassium channels mediate helium-induced preconditioning against myocardial infarction in vivo. J. Cardiothorac. Vasc. Anesth. 22:554–9.CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Huhn R, et al. (2009) Helium-induced late preconditioning in the rat heart in vivo. Br. J. Anaesth. 102:614–19.CrossRefPubMedGoogle Scholar
  21. 21.
    Oei GT, et al. (2012) Helium-induced cardioprotection of healthy and hypertensive rat myocardium in vivo. Eur. J. Pharmacol. 684:125–31.CrossRefPubMedGoogle Scholar
  22. 22.
    Oei GT, Weber NC, Hollmann MW, Preckel B. (2010) Cellular effects of helium in different organs. Anesthesiology. 112:1503–10.CrossRefPubMedGoogle Scholar
  23. 23.
    Institute of Laboratory Animal Resources; Commission on Life Sciences; National Research Council. (1996) Guide for the Care and Use of Laboratory Animals. Washington (DC): National Academy Press. See this URL for an electronic version: Scholar
  24. 24.
    Ruijter JM, et al. (2009) Amplification efficiency: linking baseline and bias in the analysis of quantitative PCR data. Nucleic Acids Res. 37:e45.CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Battke F, Symons S, Nieselt K. (2010) Mayday—integrative analytics for expression data. BMC Bioinformatics 11:121.CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Oei GT, Hollmann MW, Preckel B, Weber NC. (2012) Cardioprotection after a short episode of 70% helium inhalation at early reperfusion is abrogated by prolonged inhalation during reperfusion. Session presented at: 2012 American Society of Anesthesiologists annual meeting; 2013 Oct 13–17; Washington, DC. Abstract No. A151.Google Scholar
  27. 27.
    Hale SL, Vanderipe DR, Kloner RA. (2013) Continuous heliox breathing and the extent of anatomic zone of noreflow and necrosis following ischemia/reperfusion in the rabbit heart. Open Cardiovasc. Med. J. 8:1–5.CrossRefPubMedGoogle Scholar
  28. 28.
    Lucchinetti E, et al. (2009) Helium breathing provides modest antiinflammatory, but no endothelial protection against ischemia-reperfusion injury in humans in vivo. Anesth. Analg. 109:101–8.CrossRefPubMedGoogle Scholar
  29. 29.
    Kajstura J, et al. (1996) Apoptotic and necrotic myocyte cell deaths are independent contributing variables of infarct size in rats. Lab. Invest. 74:86–107.PubMedGoogle Scholar
  30. 30.
    Whelan RS, Kaplinskiy V, Kitsis RN. (2010) Cell death in the pathogenesis of heart disease: mechanisms and significance. Annu. Rev. Physiol. 72:19–44.CrossRefPubMedGoogle Scholar
  31. 31.
    Gao L, et al. (2012) Inhibition of autophagy contributes to ischemic postconditioning-induced neuroprotection against focal cerebral ischemia in rats. PLoS One. 7:e46092.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Fimia GM, Piacentini M. (2010) Regulation of autophagy in mammals and its interplay with apoptosis. Cell. Mol. Life Sci. 67:1581–8.CrossRefPubMedGoogle Scholar
  33. 33.
    Qiao S, et al. (2013) Delayed anesthetic preconditioning protects against myocardial infarction via activation of nuclear factor-κB and upregulation of autophagy. J. Anesth. 27:251–60.CrossRefPubMedGoogle Scholar
  34. 34.
    Wei C, Li H, Han L, Zhang L, Yang X. (2013) Activation of autophagy in ischemic postconditioning contributes to cardioprotective effects against ischemia/reperfusion injury in rat hearts. J. Cardiovasc. Pharmacol. 61:416–22.CrossRefPubMedGoogle Scholar
  35. 35.
    Lu X, Moore PG, Liu H, Schaefer S. (2011) Phosphorylation of ARC is a critical element in the antiapoptotic effect of anesthetic preconditioning. Anesth. Analg. 112:525–31.CrossRefPubMedGoogle Scholar
  36. 36.
    Matsui Y, et al. (2007) Distinct roles of autophagy in the heart during ischemia and reperfusion: roles of AMP-activated protein kinase and Beclin 1 in mediating autophagy. Circ. Res. 100:914–22.CrossRefPubMedGoogle Scholar
  37. 37.
    Takagi H, Matsui Y, Sadoshima J. (2007) The role of autophagy in mediating cell survival and death during ischemia and reperfusion in the heart. Antioxid. Redox. Signal. 9:1373–81.CrossRefPubMedGoogle Scholar
  38. 38.
    Kang R, Zeh HJ, Lotze MT, Tang D. (2011) The Beclin 1 network regulates autophagy and apoptosis. Cell Death Differ. 18:571–80.CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Klionsky DJ, et al. (2012) Guidelines for the use and interpretation of assays for monitoring autophagy. Autophagy. 8:445–544.CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    Hamasaki M, et al. (2013) Autophagosomes form at ER-mitochondria contact sites. Nature. 495:389–93.CrossRefPubMedGoogle Scholar
  41. 41.
    Gottlieb RA, Mentzer RM. (2010) Autophagy during cardiac stress: joys and frustrations of autophagy. Annu. Rev. Physiol. 72:45–59.CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    Komatsu M, Kageyama S, Ichimura Y. (2012) p62/SQSTM1/A170: physiology and pathology. Pharmacol. Res. 66:457–62.CrossRefPubMedGoogle Scholar
  43. 43.
    Pankiv S, et al. (2007) p62/SQSTM1 binds directly to Atg8/LC3 to facilitate degradation of ubiquitinated protein aggregates by autophagy. J. Biol. Chem. 282:24131–45.CrossRefPubMedGoogle Scholar
  44. 44.
    Wohlgemuth SE, Calvani R, Marzetti E. (2014) The interplay between autophagy and mitochondrial dysfunction in oxidative stress-induced cardiac aging and pathology. J. Mol. Cell. Cardiol. 71:62–70.CrossRefPubMedGoogle Scholar
  45. 45.
    Redmann M, Dodson M, Boyer-Guittaut M, Darley-Usmar V, Zhang J. (2014) Mitophagy mechanisms and role in human diseases. Int. J. Biochem. Cell. Biol. 53C:127–33.CrossRefGoogle Scholar
  46. 46.
    Huang C, et al. (2011) Preconditioning involves selective mitophagy mediated by Parkin and p62/SQSTM1. PLoS One. 6:e20975.CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Malnic B, Godfrey PA, Buck LB. (2004) Correction for Malnic et al., The human olfactory receptor gene family, PNAS 2004 101:2584–2589. Proc. Natl. Acad. Sci. U. S. A. 101:7205.CrossRefPubMedPubMedCentralGoogle Scholar
  48. 48.
    Drutel G, et al. (1995) Cloning of OL1, a putative olfactory receptor and its expression in the developing rat heart. Receptors Channels. 3:33–40.PubMedGoogle Scholar
  49. 49.
    Quach TT, et al. (2008) CRMP3 is required for hippocampal CA1 dendritic organization and plasticity. FASEB J. 22:401–9.CrossRefPubMedGoogle Scholar
  50. 50.
    Aylsworth A, Jiang SX, Desbois A, Hou ST. (2009) Characterization of the role of full-length CRMP3 and its calpain-cleaved product in inhibiting microtubule polymerization and neurite outgrowth. Exp. Cell Res. 315:2856–68.CrossRefPubMedGoogle Scholar
  51. 51.
    Winkel K, Alsheimer M, Ollinger R, Benavente R. (2009) Protein SYCP2 provides a link between transverse filaments and lateral elements of mammalian synaptonemal complexes. Chromosoma. 118:259–67.CrossRefPubMedGoogle Scholar
  52. 52.
    Simeoni F, Divita G. (2007) The Dim protein family: from structure to splicing. Cell. Mol. Life Sci. 64:2079–89.CrossRefPubMedGoogle Scholar
  53. 53.
    Dworakowski R, Alom-Ruiz SP, Shah AM. (2008) NADPH oxidase-derived reactive oxygen species in the regulation of endothelial phenotype. Pharmacol. Rep. 60:21–8.PubMedGoogle Scholar
  54. 54.
    Fukui T, et al. (2001) Expression of p22-phox and gp91-phox, essential components of NADPH oxidase, increases after myocardial infarction. Biochem. Biophys. Res. Commun. 281:1200–6.CrossRefPubMedGoogle Scholar
  55. 55.
    Chen JX, et al. (2007) NADPH oxidase modulates myocardial Akt, ERK1/2 activation, and angiogenesis after hypoxia-reoxygenation. Am. J. Physiol. Heart Circ. Physiol. 292:H1664–74.CrossRefPubMedPubMedCentralGoogle Scholar
  56. 56.
    Maejima Y, Kuroda J, Matsushima S, Ago T, Sadoshima J. (2011) Regulation of myocardial growth and death by NADPH oxidase. J. Mol. Cell. Cardiol. 50:408–16.CrossRefPubMedPubMedCentralGoogle Scholar
  57. 57.
    Bedard K, Krause KH. (2007) The NOX family of ROS-generating NADPH oxidases: physiology and pathophysiology. Physiol. Rev. 87:245–313.CrossRefPubMedGoogle Scholar
  58. 58.
    Wei CL, et al. (2011) Does conductance catheter measurement system give consistent and reliable pressure-volume relations in rats? IEEE Trans. Biomed. Eng. 58:1804–13.CrossRefPubMedGoogle Scholar
  59. 59.
    Pacher P, Nagayama T, Mukhopadhyay P, Bátkai S, Kass DA. (2008) Measurement of cardiac function using pressure-volume conductance catheter technique in mice and rats. Nat. Protoc. 3:1422–34.CrossRefPubMedPubMedCentralGoogle Scholar
  60. 60.
    Pifarré R, Cox WD, Jasuja M, Neville WE. (1969) Helium in the prevention of ventricular fibrillation. Dis. Chest. 56:135–8.CrossRefPubMedGoogle Scholar

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Authors and Affiliations

  • Gezina T. M. L. Oei
    • 1
  • Michal Heger
    • 2
  • Rowan F. van Golen
    • 2
  • Lindy K. Alles
    • 2
  • Moritz Flick
    • 1
  • Allard C. van der Wal
    • 3
  • Thomas M. van Gulik
    • 2
  • Markus W. Hollmann
    • 1
  • Benedikt Preckel
    • 1
  • Nina C. Weber
    • 1
  1. 1.Department of Anesthesiology, Laboratory of Experimental Intensive Care and Anesthesiology (L.E.I.C.A.), Academic Medical CenterUniversity of AmsterdamAmsterdamThe Netherlands
  2. 2.Department of Experimental Surgery, Academic Medical CenterUniversity of AmsterdamAmsterdamThe Netherlands
  3. 3.Department of Pathology, Academic Medical CenterUniversity of AmsterdamAmsterdamThe Netherlands

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