Electro-hyperthermia up-regulates tumour suppressor Septin 4 to induce apoptotic cell death in hepatocellular carcinoma.
Modulated electro-hyperthermia (mEHT) has been shown to be effective against various types of human tumours, including hepatocellular carcinoma (HCC). Here we aimed to investigate the molecular mechanism underlying the cytotoxic effects of mEHT to HCC cells.



Human liver cancer cell lines, Huh7 and HepG2, were treated with mEHT (42 °C/60 min) three times at 2-day intervals. Growth inhibition and apoptotic induction were evaluated using MTS, microscopic analysis, a clonogenic assay, annexin V/PI staining and a ccK18 ELISA. Global changes in gene expression were examined using RNA sequencing to obtain insights into molecular changes in response to mEHT. For in vivo evaluation of mEHT we used HepG2 HCC xenografts grown in nude mice.



mEHT suppressed HCC cell proliferation and long-term colony formation through induction of apoptosis. The growth inhibitory effects are induced through a subset of molecular changes. Notably the expression level of septin 4 (SEPT4) (involved in pro-apoptotic activity and growth suppression) was up-regulated, whereas a key regulator of invasiveness G-Protein coupled receptor 64 (GPR64) was repressed. Subsequent Western blotting confirmed that the common increase in tumour suppressor SEPT4 in both Huh7 and HepG2 cells is accompanied by the restoration of cyclin-dependent kinase (CDK) inhibitor p21 and decrease in pro-caspase 7 and pro-caspase 3, thereby accelerating apoptotic signalling in HCC cells. Additionally, mEHT significantly inhibited the growth of human HCC xenografts in nude mice.



These findings suggest that apoptotic cell death induced by mEHT is mediated by the up-regulation of tumour suppressor SEPT4 in human HCC cells.
a Department of Radiation Oncology , Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine , Seoul ;|d Department of Molecular Medicine , Keimyung University School of Medicine , Daegu , Republic of Korea.
Tae-Won
TW
Jeon
a Department of Radiation Oncology , Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine , Seoul ;
Heebum
H
Yang
a Department of Radiation Oncology , Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine , Seoul ;
Chang Geol
CG
Lee
a Department of Radiation Oncology , Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine , Seoul ;
Sang Taek
ST
Oh
b Severance Biomedical Science Institute, Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine , Seoul ;
Daekwan
D
Seo
a Department of Radiation Oncology , Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine , Seoul ;|d Department of Molecular Medicine , Keimyung University School of Medicine , Daegu , Republic of Korea.
In Hye
IH
Baik
a Department of Radiation Oncology , Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine , Seoul ;
Eun Hye
EH
Lee
a Department of Radiation Oncology , Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine , Seoul ;
Ina
I
Yun
c Department of Radiation Oncology , Ewha Women's University Medical Center , Seoul , Republic of Korea ;
Kyung Ran
KR
Park
d Department of Molecular Medicine , Keimyung University School of Medicine , Daegu , Republic of Korea.
Yun-Han
YH
Lee
D000818:Animals; D017209:Apoptosis; D006528:Carcinoma, Hepatocellular / Q000378:metabolism* / Q000473:pathology / Q000628:therapy*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D006801:Humans; D006979:Hyperthermia, Induced; D008113:Liver Neoplasms / Q000378:metabolism* / Q000473:pathology / Q000628:therapy*; D008819:Mice, Nude; D058112:Septins / Q000378:metabolism*; D047368:Tumor Burden; D015854:Up-Regulation
C492027:SEPTIN4 protein, human; D058112:Septins
Apoptosis; HCC; SEPT4; electro-hyperthermia; p21