, Volume 12, Issue 9, pp 1721–1731 | Cite as

Nanosecond pulsed electric fields induce apoptosis in p53-wildtype and p53-null HCT116 colon carcinoma cells

  • Emily H. Hall
  • Karl H. Schoenbach
  • Stephen J. BeebeEmail author
Original Paper


Non-ionizing radiation produced by nanosecond pulsed electric fields (nsPEFs) is an alternative to ionizing radiation for cancer treatment. NsPEFs are high power, low energy (non-thermal) pulses that, unlike plasma membrane electroporation, modulate intracellular structures and functions. To determine functions for p53 in nsPEF-induced apoptosis, HCT116p53+/+ and HCT116p53−/− colon carcinoma cells were exposed to multiple pulses of 60 kV/cm with either 60 ns or 300 ns durations and analyzed for apoptotic markers. Several apoptosis markers were observed including cell shrinkage and increased percentages of cells positive for cytochrome c, active caspases, fragmented DNA, and Bax, but not Bcl-2. Unlike nsPEF-induced apoptosis in Jurkat cells (Beebe et al. 2003a) active caspases were observed before increases in cytochrome c, which occurred in the presence and absence of Bax. Cell shrinkage occurred only in cells with increased levels of Bax or cytochrome c. NsPEFs induced apoptosis equally in HCT116p53+/+ and HCT116p53−/− cells. These results demonstrate that non-ionizing radiation produced by nsPEFs can act as a non-ligand agonist with therapeutic potential to induce apoptosis utilizing mitochondrial-independent mechanisms in HCT116 cells that lead to caspase activation and cell death in the presence or absence of p-53 and Bax.


Non-ionizing radiation Apoptosis Caspases Cytochrome c Bcl-2 family p53 


  1. 1.
    Beebe SJ, Fox PM, Rec LJ, Buescher ES, Somers K, Schoenbach KH (2002) Nanosecond Pulsed Electric Field (nspef) effects on cells and tissues: apoptosis induction and tumor growth inhibition. IEEE Trans Plasma Sci 30:286–292CrossRefGoogle Scholar
  2. 2.
    Beebe SJ, Fox PM, Rec LJ, Willis LK, Schoenbach KH (2003a) Nanosecond, High intensity Pulsed Electric Fields induce apoptosis in human cells. FASEB J. 17:1493–1495PubMedGoogle Scholar
  3. 3.
    Beebe SJ, White J, Blackmore PF, Deng Y, Somers K, Schoenbach KH (2003b) Diverse effects of nanosecond pulsed electric fields on cells and tissues. DNA Cell Biol 22:785–796PubMedCrossRefGoogle Scholar
  4. 4.
    Vernier PT, Sun Y, Marcu L, Craft CM, Gundersen MA (2004) Nanoelectropulse-induced phosphatidylserine translocation. Biophys J 86:4040–4048PubMedCrossRefGoogle Scholar
  5. 5.
    Weaver JC (2000) Electroporation of cells and tissues. IEEE Trans Plasma Sci 28(1):24–33CrossRefGoogle Scholar
  6. 6.
    Neumann E, Kakorin S et al (1999) Fundamentals of electroporative delivery of drugs and genes. Bioelectrochem Bioenerg 48(1):3–16PubMedCrossRefGoogle Scholar
  7. 7.
    Weaver JC, Vaughan TE et al (1999) Theory of electrical creation of aqueous pathways across skin transport barriers. Adv Drug Deliv Rev 35(1):21–39PubMedCrossRefGoogle Scholar
  8. 8.
    Heller R, Jaroszeski MJ et al (1996) Phase I/II trial for the treatment of cutaneous and subcutaneous tumors using electrochemotherapy. Cancer 77(5):964–971PubMedCrossRefGoogle Scholar
  9. 9.
    Mir LM, Morsli N, Garbay JR, Billard V, Robert C, Marty M (2003) Electrochemotherapy: a new treatment of solid tumors. J Exp Clin Cancer Res 22:145–148PubMedGoogle Scholar
  10. 10.
    Cemazar M, Golzio M, Escoffre JM, Couderc B, Sersa G, Teissie J (2006) In vivo imaging of tumor growth after electrochemotherapy with cisplatin. Biochem Biophys Res Commun 348:997–1002PubMedCrossRefGoogle Scholar
  11. 11.
    Beebe SJ, Blackmore PF, White J, Joshi RP, Schoenbach KH (2004) Nanosecond pulsed electric fields modulate cell function through intracellular signal transduction mechanisms. Physiol Meas 25:1077–1093PubMedCrossRefGoogle Scholar
  12. 12.
    Nuccitelli R Pliquett U, Chen X, Ford W, Swanson JR, Beebe SJ et al (2006) Nanosecond pulsed electric fields cause melanomas to self-destruct. Biochem Biophys Res Commun 343:351–360CrossRefGoogle Scholar
  13. 13.
    Schoenbach KH, Joshi RP, Kolb J, Chen N, Stacey M, Blackmore PF et al (2004) Ultrashort electrical pulses open a new gateway into biological cells. Proc IEEE 92:1122–1137CrossRefGoogle Scholar
  14. 14.
    Frey W, White JA, Price RO, Blackmore PF, Joshi RP, Nuccitelli R et al (2006) Plasma membrane voltage changes during Nanosecond Pulsed Electric Field exposure. Biophys J 90:3608–3615PubMedCrossRefGoogle Scholar
  15. 15.
    Gowrishankar TR, Weaver JC (2006) Electrical behavior and pore accumulation in a multicellular model for conventional and supra-electroporation. Biochem Biophys Res Commun 349:643–653PubMedCrossRefGoogle Scholar
  16. 16.
    Hu Q, Joshi RP, Schoenbach KH (2005) Simulations of nanopore formation and phosphatidylserine externalization in lipid membranes subjected to a high-intensity, ultrashort electric pulse. Phys Rev E Stat Nonlin Soft Matter Phys 72(3 Pt 1):031902PubMedGoogle Scholar
  17. 17.
    Vernier PT, Ziegler MJ, Sun Y, Chang WV, Gundersen MA, Tieleman DP (2006) Nanopore formation and phosphatidylserine externalization in a phospholipid bilayer at high transmembrane potential. J Am Chem Soc 128:6288–6289PubMedCrossRefGoogle Scholar
  18. 18.
    Schoenbach KH, Beebe SJ, Buescher ES (2001) Intracellular effect of ultrashort electrical pulses. Bioelectromagnetics 22:440–448PubMedCrossRefGoogle Scholar
  19. 19.
    Tekle E, Oubrahim H, Dzekunov SM, Kolb JF, Schoenbach KH, Chock PB (2005) Selective field effects on intracellular vacuoles and vesicle membranes with nanosecond electric pulses. Biophys J 89:274–284PubMedCrossRefGoogle Scholar
  20. 20.
    Vernier PT, Sun Y, Marcu L, Salemi S, Craft CM, Gundersen MA (2003) Calcium bursts induced by nanosecond electric pulses. Biochem Biophys Res Commun 310:286–295PubMedCrossRefGoogle Scholar
  21. 21.
    Buescher ES, Smith RR, Schoenbach KH (2004) Submicrosecond intense pulsed electric field effects on intracellular free calcium: mechanisms and effects. IEEE Trans Plasma Sci 32:1563–1572CrossRefGoogle Scholar
  22. 22.
    White JA, Blackmore PF, Schoenbach KH, Beebe SJ (2004) Stimulation of capacitative calcium entry in HL-60 cells by nanosecond pulsed electric fields. J Biol Chem 279:22964–22972PubMedCrossRefGoogle Scholar
  23. 23.
    Vernier PT, Sun Y, Marcu L, Craft CM, Gundersen MA (2004) Nanosecond pulsed electric fields perturb membrane phospholipids in T lymphoblasts. FEBS Lett 572:103–108PubMedCrossRefGoogle Scholar
  24. 24.
    Chen N, Schoenbach KH, Kolb JF, Swanson JR, Garner AL, Yang J et al (2004) Leukemic cell intracellular responses to nanosecond electric fields. Biochem Biophys Res Commun 317:421–427PubMedCrossRefGoogle Scholar
  25. 25.
    Appella E, Anderson CW (2001) Post-translational modifications and activation of p53 by genotoxic stresses. Eur J Biochem 268(10):2764–2772PubMedCrossRefGoogle Scholar
  26. 26.
    Chipuk JE, Green DR (2003) p53′s believe it or not: lessons on transcription-independent death. J Clin Immunol 23(5):355–361PubMedCrossRefGoogle Scholar
  27. 27.
    Haupt S, Berger M, Goldberg Z, Haupt Y (2003) Apoptosis––the p53 network. J Cell Sci 116:4077–4085PubMedCrossRefGoogle Scholar
  28. 28.
    Hall EM, Schoenbach KH, Beebe SJ (2005) Nanosecond pulsed electric fields induce direct electric effects and biological effects on human colon carcinoma cells. DNA Cell Biol 24:283–291PubMedCrossRefGoogle Scholar
  29. 29.
    Bortner CD, Cidlowski JA (2003) Uncoupling cell shrinkage from apoptosis reveals that Na+ influx is required for volume loss during programmed cell death. J Biol Chem 278:39176–29184PubMedCrossRefGoogle Scholar
  30. 30.
    Stacey M, Stickley J, Fox P, Statler V, Schoenbach K, Beebe SJ et al (2003) Differential effects in cells exposed to Ultra-Short, High Intensity Electric Fields: cell survival, DNA damage, and cell cycle analysis. Mutat Res 5421–2:65–75Google Scholar
  31. 31.
    Vernier PT, Sun Y, Gundersen MA (2006) Nanoelectropulse-driven membrane perturbation and small molecule permeabilization. BMC Cell Biol 19:7:37Google Scholar
  32. 32.
    Joshi RP, Hu Q, Schoenbach KH, Beebe SJ (2004) Energy-landscape-model analysis for irreversibility and its pulse-width dependence in cells subjected to a high-intensity ultrashort electric pulse. Phys Rev E Stat Nonlin Soft Matter Phys 69(5 Pt 1):051901PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2007

Authors and Affiliations

  • Emily H. Hall
    • 1
    • 2
    • 3
  • Karl H. Schoenbach
    • 2
  • Stephen J. Beebe
    • 1
    • 2
    Email author
  1. 1.Center for Pediatric Research, Children’s Hospital of the King’s Daughters and Department of Physiological SciencesEastern Virginia Medical SchoolNorfolkUSA
  2. 2.Center for BioelectricsOld Dominion University, Eastern Virginia Medical SchoolNorfolkUSA
  3. 3.Department of MicrobiologyUniversity of VirginiaCharlottesvilleUSA

Personalised recommendations