, Volume 21, Issue 1, pp 85–95 | Cite as

Knockdown of HIF-1α and IL-8 induced apoptosis of hepatocellular carcinoma triggers apoptosis of vascular endothelial cells

  • Sung Hoon Choi
  • Jun Yong ParkEmail author
  • Wonseok Kang
  • Seung Up Kim
  • Do Young Kim
  • Sang Hoon Ahn
  • Simon Wonsang Ro
  • Kwang-Hyub Han
Original Paper


A local hypoxic microenvironment is one of the most important characteristics of solid tumors. Hypoxia inducible factor-1α (HIF-1α) and Interleukin-8 (IL-8) activate tumor survival from hypoxic-induced apoptosis in each pathway. This study aimed to evaluate whether knockdown of HIF-1α and IL-8 induced apoptosis of the hepatocellular carcinoma (HCC) and endothelial cell lines. HCC cell lines were infected with adenovirus-expressing shRNA for HIF-1α and IL-8 and maintained under hypoxic conditions (1 % O2, 24 h). The expression levels of HIF-1α and both apoptotic and growth factors were examined by real-time quantitative PCR and western blot. We also investigated apoptosis by TUNEL assay (FACS and Immunofluorescence) and measured the concentration of cytochrome C. Inhibition of HIF-1α and IL-8 up-regulated the expression of apoptotic factors while downregulating anti-apoptotic factors simultaneously. Knockdown of HIF-1α and IL-8 increased the concentration of cytochrome C and enhanced DNA fragmentation in HCC cell lines. Moreover, culture supernatant collected from the knockdown of HIF-1α and IL-8 in HCC cell lines induced apoptosis in human umbilical vein endothelial cells under hypoxia, and the expression of variable apoptotic ligand increased from HCC cell lines, time-dependently. These data suggest that adenovirus-mediated knockdown of HIF-1α and IL-8 induced apoptosis in HCC cells and triggered apoptosis of vascular endothelial cells.


Apoptosis Hypoxia inducible factor-1α Interleukin-8 Death signal ligands FACs-TUNEL Tumor xenograft model 



Hypoxia inducible factor-1 alpha




Small hairpin RNA


Terminal deoxynucleotidyl transferase dUTP nick end labeling


Small hairpin RNA




Propidium iodide


Sodium dodecyl sulfate-polyacrylamide gel electrophoresis


Adenovirus mediated shRNA


Plasmid lentiviral vector mediated siRNA


TNF-related apoptosis-inducing ligand


Tumor necrosis factor-α


Fas ligand



We are grateful to Dong-Su Jang (Medical Illustrator, Medical Research Support Section, Yonsei University College of Medicine, Seoul, Korea) for his help with the figures.

Financial support

This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF), funded by the Ministry of Education, Science, and Technology (NRF-2011-0014537).

Compliance with ethical standards

Conflict of Interest

Nothing to disclose.

Supplementary material

10495_2015_1185_MOESM1_ESM.tif (1.9 mb)
Supplement 1. Compared hypoxia and normoxia in FADD expression. HUVECs were cultured under hypoxic and normoxic conditions using conditioned media prepared from the Huh7 cell line. Under the hypoxic condition, combination knockdown resulted in a higher rate of apoptosis than under the normoxic condition. Supplementary material 1 (TIFF 1939 kb)
10495_2015_1185_MOESM2_ESM.tif (1.4 mb)
Supplement 2. Comparison of death signal ligand in Hep3B cell line. In Hep3B cells, expressions of FasL, TNF-α, and TRAIL, an apoptosis inducer, were investigated over time using ELISA. With 6 h of hypoxic stimulation, there was an increase compared to the control group, though there were no significant differences. After 24 h of stimulation, FasL expression manifested remarkable differences compared to the control group. However, TNF-α and TRAIL remain unchanged. Supplementary material 2 (TIFF 1391 kb)


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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Sung Hoon Choi
    • 1
    • 2
  • Jun Yong Park
    • 1
    • 2
    • 3
    Email author
  • Wonseok Kang
    • 1
    • 2
  • Seung Up Kim
    • 1
    • 2
    • 3
  • Do Young Kim
    • 1
    • 2
    • 3
  • Sang Hoon Ahn
    • 1
    • 2
    • 3
  • Simon Wonsang Ro
    • 1
    • 2
  • Kwang-Hyub Han
    • 1
    • 2
    • 3
  1. 1.Department of Internal MedicineYonsei University College of MedicineSeoulKorea
  2. 2.Institute of GastroenterologyYonsei University College of MedicineSeoulKorea
  3. 3.Yonsei Liver CenterYonsei University Health SystemSeoulKorea

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