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Pflügers Archiv - European Journal of Physiology

, Volume 455, Issue 5, pp 939–949 | Cite as

Reduced Ca2+ entry and suicidal death of erythrocytes in PDK1 hypomorphic mice

  • Michael Föller
  • Hasan Mahmud
  • Saisudha Koka
  • Florian LangEmail author
Signal Transduction

Abstract

The phosphoinositide-dependent kinase PDK1 is a key element in the phosphoinositol-3-kinase signalling pathway, which is involved in the regulation of ion channels, transporters, cell volume and cell survival. Eryptosis, the suicidal death of erythrocytes, is characterized by decrease in cell volume, cell membrane blebbing and phospholipids scrambling with phosphatidylserine exposure at the cell surface. Oxidative stress, osmotic shock or Cl removal trigger eryptosis by activation of Ca2+-permeable cation channels and subsequent increase in cytosolic Ca2+ activity. To explore the impact of PDK1 for erythrocyte survival, eryptosis was analysed in hypomorphic mice (pdk1 hm ) expressing only some 25% of PDK1 and in their wild-type littermates (pdk1 wt ). Cell volume was estimated from forward scatter and phosphatidylserine exposure from annexin-V binding in fluorescence activated cell sorter analysis. Forward scatter was smaller in pdk1 hm than in pdk1 wt erythrocytes. Oxidative stress (100 μM tert-butylhydroperoxide), osmotic shock (+300 mM sucrose) and Cl removal (replacement of Cl with gluconate) all decreased forward scatter and increased the percentage of annexin-V-binding erythrocytes from both pdk1 hm and pdk1 wt mice. After treatment, the forward scatter was similar in both genotypes, but the percentage of annexin-V binding was significantly smaller in pdk1 hm than in pdk1 wt erythrocytes. According to Fluo-3 fluorescence, cytosolic Ca2+ activity was significantly smaller in pdk1 hm than in pdk1 wt erythrocytes. Treatment with Ca2+-ionophore ionomycin (1 μM) was followed by an increase in annexin-V binding to similar levels in pdk1 hm and pdk1 wt erythrocytes. The experiments reveal that PDK1 deficiency is associated with decreased Ca2+ entry into erythrocytes and thus with blunted eryptotic effects of oxidative stress, osmotic shock and Cl removal.

Keywords

Cell volume Ca2+ channels Phosphatidylserine Apoptosis Eryptosis 

Notes

Acknowledgements

The PDK1 hypomorphic mice have kindly been provided by Dario Alessi, Department of Biochemistry, University of Dundee, UK. The authors acknowledge the meticulous preparation of the manuscript by Jasmin Bühringer. This study was supported by the Deutsche Forschungsgemeinschaft, Nr. La 315/4-3 and La 315/6-1.

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Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Michael Föller
    • 1
  • Hasan Mahmud
    • 1
  • Saisudha Koka
    • 1
  • Florian Lang
    • 1
    Email author
  1. 1.Department of PhysiologyUniversity of TuebingenTübingenGermany

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