Tumor Biology

, Volume 37, Issue 8, pp 10415–10426 | Cite as

Hyperthermotherapy enhances antitumor effect of 5-aminolevulinic acid-mediated sonodynamic therapy with activation of caspase-dependent apoptotic pathway in human glioma

  • Donghui Ju
  • Fumio Yamaguchi
  • Guangzhi Zhan
  • Tadashi Higuchi
  • Takayuki Asakura
  • Akio Morita
  • Hideo Orimo
  • Shaoshan Hu
Original Article


Sonodynamic therapy (SDT) has shown great potential as an approach for cancer treatment, and hyperthermotherapy (HT) is also a promising cancer therapy. Here, we investigate whether HT could improve the efficacy of SDT and to make a preliminary exploration on potential mechanism. Xenograft tumor was established in nude mice model, and SNB19 and U87MG glioma cell lines were utilized for in vitro experiment. Alamar blue assay was performed to assess cell viability. Optical microscope was used to characterize the morphology changes of the glioma cells induced by SDT and HT treatments. Apoptotic rate, mitochondrial membrane potential (MMP), and intracellular production of reactive oxygen species (ROS) were examined by flow cytometer. The cell apoptosis of tumor tissues were detected by TUNEL assay. Furthermore, the expression of apoptosis-related proteins was detected with Western blot in vitro and immunohistochemistry in vivo. SDT plus HT group could significantly reduce the cell viability with circular-cell morphological change, compared with SDT group, and cell viability was decreased depending on raise of 5-ALA concentration, ultrasound exposure time, and temperature. The results also indicate that HT increased a conspicuous apoptosis, ROS production, and a remarkable loss in MMP induced by 5-ALA-SDT in vitro. Meanwhile, our data also demonstrated that the combined treatment could significantly induce apoptosis and delay tumor growth in vivo. Furthermore, in both in vitro and in vivo experiments, SDT plus HT group expressed significantly higher protein levels of Bax and cleaved caspase-3, 8, and 9 compared to SDT, HT, and control groups and significantly lower protein level of bcl-2 than the other three groups, while the expression of these proteins was unchanged between HT and control groups. HT may provide an important promotion on 5-ALA-SDT and further propose that SDT in combination with HT is a new potential application for the treatment of human glioma.


Sonodynamic therapy 5-aminolevulinic acid Hyperthermotherapy Antitumor effect Apoptosis Glioma 



We are grateful to Atsushi Chuma, Ito Co., LTD. for kindly providing ultrasonic apparatus. Miyuki Takatori and Kiyomi Kikukawa, Research Center for Life Science, Nippon Medical School, provided technical supports for the flow-cytometric analysis. We are also grateful to Ying Hu, School of Life Science and Technology, Harbin Institute of Technology, for providing technical assistance. This work was partly supported by Grant-in-Aid for challenging Exploratory Research from JSPS KAKENHI grant number 25670626 to FY. In addition, this work was also supported by grants from the Science and Technology Cooperation Project Foundation of Heilongjiang Medical Science Institute (ys14c11).

Compliance with ethical standards

Conflicts of interest



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Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2016

Authors and Affiliations

  • Donghui Ju
    • 1
    • 2
  • Fumio Yamaguchi
    • 2
  • Guangzhi Zhan
    • 2
  • Tadashi Higuchi
    • 2
  • Takayuki Asakura
    • 2
  • Akio Morita
    • 2
  • Hideo Orimo
    • 3
  • Shaoshan Hu
    • 1
  1. 1.Department of NeurosurgeryThe Second Affiliated Hospital of Harbin Medical UniversityHarbinChina
  2. 2.Department of Neurological SurgeryNippon Medical SchoolTokyoJapan
  3. 3.Division of Metabolism and Nutrition, Department of Biochemistry and Molecular BiologyNippon Medical SchoolTokyoJapan

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