Knockdown of HIF-1α and IL-8 induced apoptosis of hepatocellular carcinoma triggers apoptosis of vascular endothelial cells
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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.
KeywordsApoptosis 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
Sodium dodecyl sulfate-polyacrylamide gel electrophoresis
Adenovirus mediated shRNA
Plasmid lentiviral vector mediated siRNA
TNF-related apoptosis-inducing ligand
Tumor necrosis factor-α
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.
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.