These authors contributed equally to this work.
Hypoxia inducible factor 2 alpha inhibits hepatocellular carcinoma growth through the transcription factor dimerization partner 3/ E2F transcription factor 1–dependent apoptotic pathway†
Version of Record online: 7 FEB 2013
Copyright © 2012 American Association for the Study of Liver Diseases
Volume 57, Issue 3, pages 1088–1097, March 2013
How to Cite
Sun, H.-X., Xu, Y., Yang, X.-R., Wang, W.-M., Bai, H., Shi, R.-Y., Nayar, S. K., Devbhandari, R. P., He, Y.-z., Zhu, Q.-F., Sun, Y.-F., Hu, B., Khan, M., Anders, R. A. and Fan, J. (2013), Hypoxia inducible factor 2 alpha inhibits hepatocellular carcinoma growth through the transcription factor dimerization partner 3/ E2F transcription factor 1–dependent apoptotic pathway. Hepatology, 57: 1088–1097. doi: 10.1002/hep.26188
Potential conflict of interest: Nothing to report.
- Issue online: 28 FEB 2013
- Version of Record online: 7 FEB 2013
- Accepted manuscript online: 5 DEC 2012 12:00AM EST
- Manuscript Accepted: 12 OCT 2012
- Manuscript Revised: 11 OCT 2012
- Manuscript Received: 28 MAY 2012
- National Institutes of Health. Grant Numbers: R01DK080736, R01DK081417
- Michael Rolfe Foundation for Pancreatic Cancer Research
- Major Program of the National Natural Science Foundation of China. Grant Number: no.: 81030038
- National Key Sci-Tech Project. Grant Numbers: 2012ZX10002011-002, 2013ZX10002011-004, 2012ZX0930100-007, 2012ZX10002013-005
- National Natural Science Foundation of China. Grant Numbers: nos.: 81071661, 81000927
- Shanghai New Project for Excellent Youth. Grant Number: no.: XYQ2011020
- Zhongshan Foundation for Youth. Grant Number: no.: 2012ZSQN-06
- Research Fund for the Doctoral Program of Higher Education of China. Grant Number: no.: 20100071120064
Additional Supporting Information may be found in the online version of this article.
|HEP_26188_sm_SuppFig1.tif||608K||Supporting Information Figure 1. Expression of HIF-2α and its role in liver cancer cell. (A) The proliferation capability of MHCC97H cells cultured under normoxic or hypoxic (1% O2) conditions. (B) Expression of HIFs in MHCC97H and SMMC-7721 cells in different oxygen concentrations. (C) Expression of HIF-1α in MHCC97H clones with differing HIF-2α levels. (D) SMMC-7721 cells were transfected with pcDNA3.1-HIF-2α, pT2sh-HIF-2α, or the corresponding empty vector, then selected for G418 or puromycin resistance to isolate stably transfected clones. Left graph showed HIF-2α mRNA levels in representative monoclonal cells, error bars indicated standard deviation (n=3,* p <.01, two-tailed test). Right image: HIF-2α protein levels in representative monoclonal cells. (E) Cell proliferation as measured by CCK-8 in SMMC-7721 representative monoclonal cells. Results were expressed as the relative absorbance normalized to the one measured at zero time point. Error bars indicated standard deviation (n=6,*p <.01, two-tailed test). (F) The gene-silencing effect of siRNA targeting HIF-1α. Error bars indicate standard deviations. (G) Proliferation assay of cells with different HIF-1α or/and HIF-2α levels, cultured under 1% O2 condition. In the figure, si denotes siRNA 7146 targeting HIF-1α, and control cells were transfected with siRNA containing a scrambled sequence. Error bars indicate standard deviations. *p <0.05. (H) Cell proliferation measured by Ki67 in MHCC97H clone. Quantification of the relative percentage of Ki67 positive cells were shown in right image. Error bars indicated standard deviation (n=6, two-tailed test).|
|HEP_26188_sm_SuppFig2.tif||1298K||Supporting Information Figure 2. HIF-2α expression in orthotropic xenografts. (A) HIF-2α mRNA expression was confirmed in tumors formed by MHCC97H monoclones. (B) CD31 staining in tumors.|
|HEP_26188_sm_SuppFig3.tif||2427K||Supporting Information Figure 3. HIF-2α augments apoptosis by inhibiting the expression of TFDP3 Classification of HIF-2α-bound genes identified by ChIP-on-chip assay based on the biological function according to the enrichment score. It should be noted that some genes may be represented in more than one classified group.(B) HIF-2α target genes identified by ChIP-on-chip were classified based on cellular component according to the enrichment score. (C) HIF-1α levels in MHCC97H cells, which were transiently transfected by siRNA, pcDNA3-HIF-2α expression vectors, or control sequences. (D) TFDP3 levels were measured in MHCC97H cells with different HIF-1α levels. (E) The RNA level of TFDP3 and HIF-2α were tested in MHCC97H cells which was transiently transfected with pcDNA3.1- HIF-2α, pcDNA3.1-TFDP3 or mock plasmid. (F) The mRNA levels of other members of DP family were confirmed in the MHCC97H cells which were transiently transfected with HIF-2α (pcDNA3.1- HIF-2α), TFDP3 (pcDNA3.1-TFDP3) or empty vector (pcDNA3.1). (G) Gene-silencing efficiency of siRNA targeting E2F1. Error bars indicate standard deviations. (H) Proliferation of HIF-2α-overexpressing MHCC97H clones with different levels of E2F1. Error bars indicate standard deviations. *p<0.05. (I) The expression levels of apoptosis genes in HIF-2α-overexpressing MHCC97H cells with different levels of E2F1.|
|HEP_26188_sm_SuppTabs.doc||126K||Supporting Information Tables|
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