Effect of Low-Intensity Ultrasound on Mortality of PC12 Induced by Amyloid β25–35

  • Chun-Yi ChiuEmail author
  • Shyh-Hau Wang
Original Article


The aim of this study is to investigate the potential of using ultrasound (US) to protect neuronal cells from damage by amyloid beta (Αβ) peptide. Experiments were performed using PC12 cells with the addition of 20 µM Aβ25–35, US stimulation, or both. 1-MHz US at a 20 % duty cycle with various intensities (10, 50, 10, and 150 mW/cm2) for 3 min was employed. The responses of PC12 cells were determined in terms of the survival rate via the MTT assay, cell morphology via optical microscopy, and cell apoptosis/necrosis characteristics via Annexin V-fluorescein isothiocyanate (FITC) and propidium iodide (PI) fluorescence staining and terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick-end-labeling assay. The results show that the survival rate of PC12 cells in the presence of Aβ23–35 increased by 4.8–6 % when the cells were exposed to 100- and 150-mW/cm2 US. With US intensity of 150 mW/cm2, the growth of neurites from PC12 cells was observed. The experiment results of cell stains with Annexin V-FITC/PI show that US decreased PC12 cell apoptosis and necrosis. In addition, for the PC12 cells with added Aβ25–35, the fold change in cell apoptosis decreased by 0.36 when a US intensity of 150 mW/cm2 was applied. In conclusion, low-intensity US decreases PC12 cell mortality caused by Aβ23–35.


Amyloid beta peptide PC12 cells Low-intensity ultrasound 



The authors would like to thank the Ministry of Science and Technology of the Republic of China for partially financially supporting this research under Grant National Science Council 94-2213-E-033-039.


  1. 1.
    Waring, S. C., & Rosenberg, R. N. (2008). Genome-wide association studies in Alzheimer disease. Archives of Neurology, 65, 329–334.CrossRefGoogle Scholar
  2. 2.
    Craig, L. A., Hong, N. S., & McDonald, R. J. (2011). Revisiting the cholinergic hypothesis in the development of Alzheimer’s disease. Neuroscience and Biobehavioral Reviews, 35, 1397–1409.CrossRefGoogle Scholar
  3. 3.
    Karran, E., Mercken, M., & De Strooper, B. (2011). The amyloid cascade hypothesis for Alzheimer’s disease: an appraisal for the development of therapeutics. Nature Reviews Drug Discovery, 10, 698–712.CrossRefGoogle Scholar
  4. 4.
    Maccioni, R. B., Farías, G., Morales, I., & Navarrete, L. (2010). The revitalized tau hypothesis on Alzheimer’s disease. Archives of Medical Research, 41, 226–231.CrossRefGoogle Scholar
  5. 5.
    Kawahara, M., & Kuroda, Y. (2000). Molecular mechanism of neurodegeneration induced by Alzheimer’s beta-amyloid protein: channel formation and disruption of calcium homeostasis. Brain Research Bulletin, 53, 389–397.CrossRefGoogle Scholar
  6. 6.
    Polinsky, R. J. (1998). Clinical pharmacology of rivastigmine: a new-generation acetylcholinesterase inhibitor for the treatment of Alzheimer’s disease. Clinical Therapeutics, 20, 634–647.CrossRefGoogle Scholar
  7. 7.
    Wilkinson, D., & Murray, J. (2001). Galantamine: a randomized, double-blind, dose comparison in patients with Alzheimer’s disease. International Journal of Geriatric Psychiatry, 16, 852–857.CrossRefGoogle Scholar
  8. 8.
    Feldman, H., Gauthier, S., Hecker, J., Vellas, B., Subbiah, P., Whalen, E., & D. M. S. I. Group. (2001). A 24-week, randomized, double-blind study of donepezil in moderate to severe Alzheimer’s disease. Neurology, 57, 613–620.CrossRefGoogle Scholar
  9. 9.
    Knopman, D. S., & Morris, J. C. (1997). An update on primary drug therapies for Alzheimer disease. Archives of Neurology, 54, 1406–1409.CrossRefGoogle Scholar
  10. 10.
    Farlow, M. R., & Evans, R. M. (1998). Pharmacologic treatment of cognition in Alzheimer’s dementia. Neurology, 51, S36–S44.CrossRefGoogle Scholar
  11. 11.
    Peskind, E. R., Potkin, S. G., Pomara, N., Ott, B. R., Graham, S. M., Olin, J. T., & McDonald, S. (2006). Memantine treatment in mild to moderate Alzheimer disease: a 24-week randomized, controlled trial. The American Journal of Geriatric Psychiatry, 14, 704–715.CrossRefGoogle Scholar
  12. 12.
    Schneider, L. S., Dagerman, K. S., Higgins, J. P., & McShane, R. (2011). Lack of evidence for the efficacy of memantine in mild Alzheimer disease. Archives of Neurology, 68, 991–998.CrossRefGoogle Scholar
  13. 13.
    Lim, G. P., Yang, F., Chu, T., Chen, P., Beech, W., Teter, B., et al. (2000). Ibuprofen suppresses plaque pathology and inflammation in a mouse model for Alzheimer’s disease. Journal of Neuroscience, 20, 5709–5714.Google Scholar
  14. 14.
    Lovell, M. A., Xie, C., & Markesbery, W. R. (1999). Protection against amyloid beta peptide toxicity by zinc. Brain Research, 823, 88–95.CrossRefGoogle Scholar
  15. 15.
    Alvarez, G., Muñoz-Montaño, J. R., Satrústegui, J., Avila, J., Bogónez, E., & Díaz-Nido, J. (1999). Lithium protects cultured neurons against beta-amyloid-induced neurodegeneration. FEBS Letters, 453, 260–264.CrossRefGoogle Scholar
  16. 16.
    Hosoda, T., Nakajima, H., & Honjo, H. (2001). Estrogen protects neuronal cells from amyloid beta-induced apoptotic cell death. Neuroreport, 12, 1965–1970.CrossRefGoogle Scholar
  17. 17.
    Grundman, M. (2000). Vitamin E and Alzheimer disease: the basis for additional clinical trials. The American Journal of Clinical Nutrition, 71, 630S–636S.Google Scholar
  18. 18.
    Luchsinger, J. A., Tang, M. X., Shea, S., & Mayeux, R. (2003). Antioxidant vitamin intake and risk of Alzheimer disease. Archives of Neurology, 60, 203–208.CrossRefGoogle Scholar
  19. 19.
    Pardridge, W. M. (2009). Alzheimer’s disease drug development and the problem of the blood-brain barrier. Alzheimers Dement, 5, 427–432.CrossRefGoogle Scholar
  20. 20.
    Banks, W. A. (2012). Drug delivery to the brain in Alzheimer’s disease: consideration of the blood-brain barrier. Advanced Drug Delivery Reviews, 64, 629–639.CrossRefGoogle Scholar
  21. 21.
    Scherder, E. J., Bouma, A., & Steen, A. M. (1995). Effects of short-term transcutaneous electrical nerve stimulation on memory and affective behaviour in patients with probable Alzheimer’s disease. Behavioural Brain Research, 67, 211–219.CrossRefGoogle Scholar
  22. 22.
    Scherder, E. J., & Bouma, A. (1999). Effects of transcutaneous electrical nerve stimulation on memory and behavior in Alzheimer’s disease may be stage-dependent. Biological Psychiatry, 45, 743–749.CrossRefGoogle Scholar
  23. 23.
    Scherder, E. J., Van Someren, E. J., & Swaab, D. F. (1999). Transcutaneous electrical nerve stimulation (TENS) improves the rest-activity rhythm in midstage Alzheimer’s disease. Behavioural Brain Research, 101, 105–107.CrossRefGoogle Scholar
  24. 24.
    Duan, R., Zhu, L., Liu, T. C., Li, Y., Liu, J., Jiao, J., et al. (2003). Light emitting diode irradiation protect against the amyloid beta 25–35 induced apoptosis of PC12 cell in vitro. Lasers in Surgery and Medicine, 33, 199–203.CrossRefGoogle Scholar
  25. 25.
    Zhang, L., Xing, D., Zhu, D., & Chen, Q. (2008). Low-power laser irradiation inhibiting Abeta25–35-induced PC12 cell apoptosis via PKC activation. Cellular Physiology and Biochemistry, 22, 215–222.CrossRefGoogle Scholar
  26. 26.
    Crisci, A. R., & Ferreira, A. L. (2002). Low-intensity pulsed ultrasound accelerates the regeneration of the sciatic nerve after neurotomy in rats. Ultrasound in Medicine and Biology, 28, 1335–1341.CrossRefGoogle Scholar
  27. 27.
    Ter Haar, G. (1999). Therapeutic ultrasound. European Journal of Ultrasound, 9, 3–9.CrossRefGoogle Scholar
  28. 28.
    Ward, J. F. (2011). High-intensity focused ultrasound for therapeutic tissue ablation in surgical oncology. Surgical Oncology Clinics of North America, 20, 389–407.CrossRefGoogle Scholar
  29. 29.
    Yoshizawa, S., Ikeda, T., Ito, A., Ota, R., Takagi, S., & Matsumoto, Y. (2009). High intensity focused ultrasound lithotripsy with cavitating microbubbles. Medical & Biological Engineering & Computing, 47, 851–860.CrossRefGoogle Scholar
  30. 30.
    Ter Haar, G. (2007). Therapeutic applications of ultrasound. Progress in Biophysics and Molecular Biology, 93, 111–129.CrossRefGoogle Scholar
  31. 31.
    Sena, K., Leven, R. M., Mazhar, K., Sumner, D. R., & Virdi, A. S. (2005). Early gene response to low-intensity pulsed ultrasound in rat osteoblastic cells. Ultrasound in Medicine and Biology, 31, 703–708.CrossRefzbMATHGoogle Scholar
  32. 32.
    Chen, S., Wu, C., Wang. S., & Li, W. (2014). Growth and differentiation of osteoblasts regulated by low-intensity pulsed ultrasound of various exposure durations. Journal of Medical and Biological Engineering, 34, 197–203.Google Scholar
  33. 33.
    Mukai, S., Ito, H., Nakagawa, Y., Akiyama, H., Miyamoto, M., & Nakamura, T. (2005). Transforming growth factor-beta1 mediates the effects of low-intensity pulsed ultrasound in chondrocytes. Ultrasound in Medicine and Biology, 31, 1713–1721.CrossRefGoogle Scholar
  34. 34.
    Breuing, K., Bayer, L., Neuwalder, J., & Orgill, D., (2005). Early experience using low-frequency ultrasound in chronic wounds. Annals of Plastic Surgery, 55, 183–187.Google Scholar
  35. 35.
    De Deyne, P. G., & Kirsch-Volders, M. (1995). In vitro effects of therapeutic ultrasound on the nucleus of human fibroblasts. Physical Therapy, 75, 629–634.Google Scholar
  36. 36.
    Webster, D. F., Pond, J. B., Dyson, M., & Harvey, W. (1978). The role of cavitation in the in vitro stimulation of protein synthesis in human fibroblasts by ultrasound. Ultrasound in Medicine and Biology, 4, 343–351.CrossRefGoogle Scholar
  37. 37.
    Webster, D. F., Harvey, W., Dyson, M., & Pond, J. B. (1980). The role of ultrasound-induced cavitation in the ‘in vitro’ stimulation of collagen synthesis in human fibroblasts. Ultrasonics, 18, 33–37.CrossRefGoogle Scholar
  38. 38.
    Tsai, W. C., Hsu, C. C., Tang, F. T., Chou, S. W., Chen, Y. J., & Pang, J. H. (2005). Ultrasound stimulation of tendon cell proliferation and upregulation of proliferating cell nuclear antigen. Journal of Orthopaedic Research, 23, 970–976.CrossRefGoogle Scholar
  39. 39.
    Zhou, S., Schmelz, A., Seufferlein, T., Li, Y., Zhao, J., & Bachem, M. G. (2004). Molecular mechanisms of low intensity pulsed ultrasound in human skin fibroblasts. Journal of Biological Chemistry, 279, 54463–54469.CrossRefzbMATHGoogle Scholar
  40. 40.
    Parvizi, J., Parpura, V., Greenleaf, J. F., & Bolander, M. E. (2002). Calcium signaling is required for ultrasound-stimulated aggrecan synthesis by rat chondrocytes. Journal of Orthopaedic Research, 20, 51–57.CrossRefzbMATHGoogle Scholar
  41. 41.
    Ashush, H., Rozenszajn, L. A., Blass, M., Barda-Saad, M., Azimov, D., Radnay, J., et al. (2000). Apoptosis induction of human myeloid leukemic cells by ultrasound exposure. Cancer Research, 60, 1014–1020.Google Scholar
  42. 42.
    Firestein, F., Rozenszajn, L. A., Shemesh-Darvish, L., Elimelech, R., Radnay, J., & Rosenschein, U. (1010). Induction of apoptosis by ultrasound application in human malignant lymphoid cells: role of mitochondria-caspase pathway activation. Annals of the New York Academy of Sciences, 163–166, 2003.Google Scholar
  43. 43.
    O’Brien, W. D. (2007). Ultrasound-biophysics mechanisms. Progress in Biophysics and Molecular Biology, 93, 212–255.CrossRefzbMATHGoogle Scholar
  44. 44.
    Wu, J., & Nyborg, W. L. (2008). Ultrasound, cavitation bubbles and their interaction with cells. Advanced Drug Delivery Reviews, 60, 1103–1116.CrossRefGoogle Scholar
  45. 45.
    Huang, H., Kamm, R. D., & Lee, R. T. (2004). Cell mechanics and mechanotransduction: pathways, probes, and physiology. American Journal of Physiology. Cell Physiology, 287, C1–C11.CrossRefGoogle Scholar
  46. 46.
    Zhang, H., Wu, S., & Xing, D. (2012). Inhibition of Aβ(25–35)-induced cell apoptosis by low-power-laser-irradiation (LPLI) through promoting Akt-dependent YAP cytoplasmic translocation. Cellular Signalling, 24, 224–232.CrossRefGoogle Scholar
  47. 47.
    Kimura, K., Yanagida, Y., Haruyama, T., Kobatake, E., & Aizawa, M. (1998). Electrically induced neurite outgrowth of PC12 cells on the electrode surface. Medical and Biological Engineering and Computing, 36, 493–498.CrossRefGoogle Scholar
  48. 48.
    Kimura, K., Yanagida, Y., Haruyama, T., Kobatake, E., & Aizawa, M. (1998). Gene expression in the electrically stimulated differentiation of PC12 cells. Journal of Biotechnology, 63, 55–65.CrossRefzbMATHGoogle Scholar

Copyright information

© Taiwanese Society of Biomedical Engineering 2015

Authors and Affiliations

  1. 1.Department of Biomedical EngineeringChung Yuan Christian UniversityChung LiTaiwan, ROC
  2. 2.Departmant of Computer Science and Information EngineeringNational Cheng Kung UniversityTainanTaiwan, ROC

Personalised recommendations