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Microenvironment and Immunology

In Vivo Profiling of Hypoxic Gene Expression in Gliomas Using the Hypoxia Marker EF5 and Laser-capture Microdissection

Diane Marotta, Jayashree Karar, W. Timothy Jenkins, Monika Kumanova, Kevin W. Jenkins, John W. Tobias, Donald Baldwin, Artemis Hatzigeorgiou, Panagiotis Alexiou, Sydney M. Evans, Rodolfo Alarcon, Amit Maity, Cameron Koch and Constantinos Koumenis
Diane Marotta
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Jayashree Karar
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W. Timothy Jenkins
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Monika Kumanova
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Kevin W. Jenkins
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John W. Tobias
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Donald Baldwin
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Artemis Hatzigeorgiou
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Panagiotis Alexiou
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Sydney M. Evans
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Rodolfo Alarcon
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Amit Maity
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Cameron Koch
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Constantinos Koumenis
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DOI: 10.1158/0008-5472.CAN-10-3061 Published February 2011
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Abstract

Hypoxia is a key determinant of tumor aggressiveness, yet little is known regarding hypoxic global gene regulation in vivo. We used the hypoxia marker EF5 coupled with laser-capture microdissection to isolate RNA from viable hypoxic and normoxic regions of 9L experimental gliomas. Through microarray analysis, we identified several mRNAs (including the HIF targets Vegf, Glut-1, and Hsp27) with increased levels under hypoxia compared with normoxia both in vitro and in vivo. However, we also found striking differences between the global in vitro and in vivo hypoxic mRNA profiles. Intriguingly, the mRNA levels of a substantial number of immunomodulatory and DNA repair proteins including CXCL9, CD3D, and RAD51 were found to be downregulated in hypoxic areas in vivo, consistent with a protumorigenic role of hypoxia in solid tumors. Immunohistochemical staining verified increased HSP27 and decreased RAD51 protein levels in hypoxic versus normoxic tumor regions. Moreover, CD8+ T cells, which are recruited to tumors upon stimulation by CXCL9 and CXCL10, were largely excluded from viable hypoxic areas in vivo. This is the first study to analyze the influence of hypoxia on mRNA levels in vivo and can be readily adapted to obtain a comprehensive picture of hypoxic regulation of gene expression and its influence on biological functions in solid tumors. Cancer Res; 71(3); 779–89. ©2011 AACR.

Footnotes

  • Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/).

  • Received August 19, 2010.
  • Revision received November 11, 2010.
  • Accepted November 26, 2010.
  • ©2011 American Association for Cancer Research.
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Cancer Research: 71 (3)
February 2011
Volume 71, Issue 3
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In Vivo Profiling of Hypoxic Gene Expression in Gliomas Using the Hypoxia Marker EF5 and Laser-capture Microdissection
Diane Marotta, Jayashree Karar, W. Timothy Jenkins, Monika Kumanova, Kevin W. Jenkins, John W. Tobias, Donald Baldwin, Artemis Hatzigeorgiou, Panagiotis Alexiou, Sydney M. Evans, Rodolfo Alarcon, Amit Maity, Cameron Koch and Constantinos Koumenis
Cancer Res February 1 2011 (71) (3) 779-789; DOI: 10.1158/0008-5472.CAN-10-3061

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In Vivo Profiling of Hypoxic Gene Expression in Gliomas Using the Hypoxia Marker EF5 and Laser-capture Microdissection
Diane Marotta, Jayashree Karar, W. Timothy Jenkins, Monika Kumanova, Kevin W. Jenkins, John W. Tobias, Donald Baldwin, Artemis Hatzigeorgiou, Panagiotis Alexiou, Sydney M. Evans, Rodolfo Alarcon, Amit Maity, Cameron Koch and Constantinos Koumenis
Cancer Res February 1 2011 (71) (3) 779-789; DOI: 10.1158/0008-5472.CAN-10-3061
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