Metabolic correlates of tumour hypoxia in malignant canine mammary carcinoma G. Mees a , C. Vangestel b , R. Dierckx a,g , S. Loomans c , N. Van Damme b , K. Peremans d , H. De Rooster e , B. Van Goethem e , P. Pauwels f , R. Ducatelle c , C. Van de Wiele a,g,⇑ a Department of Nuclear Medicine and Molecular Imaging, University Medical Center Groningen, University of Groningen, The Netherlands b Department of Gastroenterology, Ghent University Hospital, Belgium c Department of Pathology, Faculty of Veterinary Medicine, Ghent, Belgium d Department of Medical Imaging, Faculty of Veterinary Medicine, Ghent, Belgium e Department of Small Animal Medicine and Clinical Biology, Faculty of Veterinary Medicine, Ghent, Belgium f Department of Pathology, Ghent University Hospital, Belgium g Department of Nuclear Medicine, Ghent University Hospital, Belgium article info Article history: Received 2 September 2010 Accepted 18 January 2011 Keywords: Tumour hypoxia Pimonidazole GLUT abstract Given its importance in human and canine tumour biology, a profound understanding of tumour hypoxia is of paramount importance. Therefore, the aim of this work was to investigate the relationship between tumour hypoxia and the expression of a number of hypoxia-induced proteins that play a role in tumour metabolism. The hypoxia marker pimonidazole was administered to dogs affected by spontaneous mam- mary carcinoma and compared with immunohistochemical staining for GLUT1 and 3, HK 2 and CA IX. A statistically significant correlation was found between pimonidazole staining and GLUT1-expression (R = 0.607; p = 0.001). These results indicate a strong interaction between tumour hypoxia and tumour metabolism by the induction of proteins essential to maintain a stable tumour microenvironment. Ó 2011 Elsevier Ltd. All rights reserved. 1. Introduction The occurrence of heterogeneously distributed hypoxic and/or anoxic areas is a characteristic feature of locally advanced solid tu- mours and has been described in a wide range of human as well as canine malignancies (Snyder et al., 2008). These areas of acute and chronic hypoxia are the result of an abnormal structure and func- tion of the tumoural microvessels, increased diffusion distances between the nutritive blood vessels and the tumour cells, and disease- or treatment-related anaemia. Recent studies have dem- onstrated a clear relationship between this hypoxic microenviron- ment and tumour-associated metabolic alterations. In this respect, tumour hypoxia has been associated with an aggressive tumour phenotype, poor response to therapy, and worse prognosis in a wide range of human malignancies (Höckel and Vaupel, 2001; Vaupel and Harrison, 2004; Vaupel and Mayer, 2007). Cancer cells adapt to hypoxic conditions by the induction of target genes in- volved in glucose metabolism, angiogenesis, erythropoiesis and apoptosis in an effort to overcome their compromised condition. Many of these adaptations are coordinated by the transcription factor hypoxia-inducible factor (HIF)-1, which has been verified as a master regulator of oxygen homeostasis under hypoxic condi- tions (Semenza, 2000). To compensate for the hypoxia-induced drop in mitochondrially produced ATP, cells will increase their glycolytic ATP-production (Pasteur effect) which is inherently inef- ficient when compared to aerobic degradation of glucose (Krebs cycle), leading to an increase in glucose transport and consump- tion. This is further enhanced by aerobic glycolysis (Warburg ef- fect) (Warburg, 1956). Several approaches, non-invasive and invasive, exist for the detection of tumour hypoxia (Vaupel and Mayer, 2007). Numerous studies have investigated tumour hypox- ia in a prospective and retrospective manner by studying the expression and localisation of endogenous and exogenous markers of hypoxia. Several different candidates have emerged as possible endogenous markers of hypoxia of which the most important are HIF-1a, carbonic anhydrase IX (CA IX), glucose transporter (GLUT) 1 and 3. None of these have however, proven their selectivity for hypoxia as their expression is believed to be influenced by other parameters than merely hypoxia (Bussink et al., 2003). The most relevant marker for qualitative and quantitative determination of tumour hypoxia in experimental and clinical circumstances is the exogenous hypoxia marker pimonidazole. This 2-nitroimidazole reagent is reduced under hypoxic conditions and the resulting ad- ducts are irreversibly bound to thiol-containing proteins in viable hypoxic cells (pO 2 < 10 mmHg) (Gross et al., 1995). Pimonidazole is a robust marker of hypoxia and its microregional distribution correlates with oxygen concentration at the cellular level (Raleigh 0034-5288/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.rvsc.2011.01.014 ⇑ Corresponding author at: Ghent University Hospital, Department of Nuclear Medicine, De Pintelaan 185, 9000 Ghent, Belgium. Tel.: +32 (0) 93323028; fax: +32 (0) 93323807. E-mail address: christophe.vandewiele@ugent.be (C. Van de Wiele). Research in Veterinary Science 91 (2011) e125–e128 Contents lists available at ScienceDirect Research in Veterinary Science journal homepage: www.elsevier.com/locate/rvsc