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Journal of Neuro-Oncology

, Volume 85, Issue 1, pp 49–63 | Cite as

Quantifying the A1AR distribution in peritumoural zones around experimental F98 and C6 rat brain tumours

  • Markus Dehnhardt
  • Christoph Palm
  • Andrea Vieten
  • Andreas Bauer
  • Uwe Pietrzyk
Lab. Investigation-human/animal tissue

Abstract

Quantification of growth in experimental F98 and C6 rat brain tumours was performed on 51 rat brains, 17 of which have been further assessed by 3D tumour reconstruction. Brains were cryosliced and radio-labelled with a ligand of the peripheral type benzodiazepine-receptor (pBR), 3H-Pk11195 [(1-(2-chlorophenyl)-N-methyl-N-(1-methyl-propylene)-3-isoquinoline-carboxamide)] by receptor autoradiography. Manually segmented and automatically registered tumours have been 3D-reconstructed for volumetric comparison on the basis of 3H-Pk11195-based tumour recognition. Furthermore automatically computed areas of −300 μm inner (marginal) zone as well as 300 μm and 600 μm outer tumour space were quantified. These three different regions were transferred onto other adjacent slices that had been labelled by receptor autoradiography with the A1 Adenosine receptor (A1AR)-ligand 3H-CPFPX (3H-8-cyclopentyl-3-(3-fluorpropyl)-1-propylxanthine) for quantitative assessment of A1AR in the three different tumour zones. Hence, a method is described for quantifying various receptor protein systems in the tumour as well as in the marginal invasive zones around experimentally implanted rat brain tumours and their representation in the tumour microenvironment as well as in 3D space. Furthermore, a tool for automatically reading out radio-labelled rat brain slices from auto radiographic films was developed, reconstructed into a consistent 3D-tumour model and the zones around the tumour were visualized. A1AR expression was found to depend upon the tumour volume in C6 animals, but is independent on the time of tumour development. In F98 animals, a significant increase in A1AR receptor protein was found in the Peritumoural zone as a function of time of tumour development and tumour volume.

Keywords

3D reconstruction A1 adenosine receptor GBM Kmeans algorithm Brain tumour Receptor autoradiography 

Abbreviations

A1AR

A1 adenosine receptor

C6

Cloned rat glial tumour cell line

CPFPX

8-Cyclopentyl-3-(3-fluorpropyl)-1-propylxanthine)

F98

Astrocytic tumour cell line

GBM

Glioblastoma multiforme

Pk11195

1-(2-chlorphenyl)-N-methyl-N-(1-methyl-propylene)-3-isoquinoline-carboxamide

pBR

Peripheral-type benzodiazepine receptor

R-PIA

(R(-)-N-(2-phenylisopropyl)-adenosine

References

  1. 1.
    Bauer A, Langen KJ, Bidmon H, Holschbach MH, Weber S, Olson RA, Coenen HH, Zilles K (2005) 18F-Cyclopentyl-3(3-18F-Fluoropropyl)-1-Propyl-xanthine PET identifies changes in cerebral A1Adenosine receptor density caused by glioma invasion. J Nucl Med 46:450–454PubMedGoogle Scholar
  2. 2.
    Hoelzinger DB, Mariani L, Woyke T, Berens TJ, McDonough WS, Sloan A, Coons SW, Berens M (2005) Gene expression profile of glioblastoma multiforme invasive phenotype points to new therapeutic targets. Neoplasia 7(1):7–16PubMedCrossRefGoogle Scholar
  3. 3.
    Liyang Y, Diehn M, Watson N, Bollen AW, Aldape KD, Nicholas MK, Lamborn KR, Berger MS, Botstein D, Brown PO, Israel MA (2005) Gene expression profiling reveales molecularly and clinically distinct subtypes of glioblastoma multiforme. Proc Natl Acad Sci USA 102(16):5814–5819CrossRefGoogle Scholar
  4. 4.
    Vajkoczy P, Schilling L, Ullrich A, Schmiedek P, Menger MD (1998) Characterization of angiogenesis and microcirculation of high-grade glioma: an intravital multifluorescence microscopic approach in the athymic nude mouse. J Cereb Blood Flow Metab 18:510–520PubMedCrossRefGoogle Scholar
  5. 5.
    Wesseling P, Van der Laak JAWM, De Leeuw H, Ruiter DJ, Burger PC (1994) Quantitative immunohistological analysis of the microvasculature in untreated human glioblastoma multiforme; computer-assisted image analysis on whole tumour sections. J Neurosurg 81:902–909PubMedCrossRefGoogle Scholar
  6. 6.
    Acker T, Acker H (2004) Cellular oxygen sensing need in CNS functions: physiological and pathological implications. J Exp Biol 207:3171–3188PubMedCrossRefGoogle Scholar
  7. 7.
    Borg SA, Kerry KE, Royds JA, Battersby RD, Jones TH (2005) Correlation of VEGF production with IL1a and IL6 secretion by human pituitary adenoma cells. Eur J Endocrinol 152:293–300PubMedCrossRefGoogle Scholar
  8. 8.
    Khoo HE, Ho CL, Chatwal VJS, Chan STF, Ngoi SS, Moochhala SM (1996) Differential expression of adenosine A1 receptors in colorectal cancer and related mucosa. Cancer Lett 106:17–21PubMedCrossRefGoogle Scholar
  9. 9.
    Ralevic V, Burnstock G (1998) Receptors for purines and pyrimidines. Pharm Rev 50(3):413–492PubMedGoogle Scholar
  10. 10.
    Barth RF (1998) Rat brain tumour models in experimental neuro-oncology: the 9L, C6, T9, F98, RG2 (D74), RT-2 and CNS-1 Gliomas. J Neurooncol 36:91–102PubMedCrossRefGoogle Scholar
  11. 11.
    Benda P (1968) Differentiated rat glial cell strain in tissue culture. Science 161:370–371PubMedCrossRefGoogle Scholar
  12. 12.
    Danielson PE (1980) Euclidean distance mapping. Comput Graphics Image Process 14:227–248CrossRefGoogle Scholar
  13. 13.
    Miyazawa N, Hamel E, Diksic M (1998) Assessment of the peripheral benzodiazepine receptors in human gliomas by two methods. J Neurooncol 38:19–26PubMedCrossRefGoogle Scholar
  14. 14.
    Jordà EG, Jiménez A, Verdaguer E, Canudas AM, Folch J, Sureda FX, Camins A, Pallàs M (2005) Evidence in favour of a role for peripheral-type benzodiazepine receptor ligands in amplification of neuronal apoptosis. Apoptosis 10:91–104PubMedCrossRefGoogle Scholar
  15. 15.
    Duda RO, Hart PE (1973) Pattern classification and scene analysis. J Wiley & Sons, New York, USAGoogle Scholar
  16. 16.
    Kim J, Fessler JA (2004) Intensity-based image registration using robust correlation coefficients. IEEE Trans Med Imag 23(11):1430–1444CrossRefGoogle Scholar
  17. 17.
    Ibanez L, Schroeder W, Ng L, Cates J (2005) The ITK software guide, 2nd edn. http://www.itk.orgGoogle Scholar
  18. 18.
    Paxinos G, Watson C (1986) The rat brain in stereotaxic coordinates. Academic Press, SydneyGoogle Scholar
  19. 19.
    Palm C, Dehnhardt M, Vieten A and Pietrzyk U (2005) 3D rat brain tumour reconstruction. Proceedings of 14th International Conference of IOMP, EFOMP and DGMP (ICMP 2005) and of 39th Annual Congress of DGBMT within VDE (BMT 2005). Kalender W, Hahn EG, Schulte AM (eds) Biomedizinische Technik 50(1):597–598Google Scholar
  20. 20.
    Bifulco M, DiMarzo V (2002) Targeting the endocannabinoid system in cancer therapy: a call for further research. Nat Med 8(6):547–550PubMedCrossRefGoogle Scholar
  21. 21.
    Kirsch M, Schackert G, Black PMcL (2000) Anti-angiogenic treatment strategies for malignant brain tumours. J Neurooncol 50:149–163PubMedCrossRefGoogle Scholar
  22. 22.
    Biber K, Lubrich B, Fiebich BL, Boddeke HWGM, von Calker D (2001) Interleukin-6 enhances expression of Adenosine A1 receptor mRNA and signalling in cultured rat cortical astrocytes and brain slices. Psychpharmacol 24:86–96Google Scholar
  23. 23.
    Dulak J, Józkowicz A (2005) Anti-angiogenic and anti-inflammatory effects of statins: relevance to anti-cancer therapy. Curr Cancer Drug Targets 5(8):579–594PubMedCrossRefGoogle Scholar
  24. 24.
    Zhang Y, Zhao W, Zhang HJ, Domann FE, Oberley LW (2002) Over-expression of copper zinc superoxide dismutase suppresses human glioma cell growth. Cancer Res 62:1205–1212PubMedGoogle Scholar
  25. 25.
    Sun Y, Schmidt NO, Schmidt K, Doshi S, Rubin JB, Mulkern RV, Carroll R, Ziu M, Erkmen K, Poussaint TY, Black P, Albert M, Burstein D, Kieran MW (2004) Perfusion MRI of U87 brain tumours in a mouse model. Magn Res Med 51:893–899CrossRefGoogle Scholar
  26. 26.
    Bhujwalla ZM, Artemov D, Natarajan K, Ackerstaff E, Solaiyappan M (2001) Vascular differences detected by MRI for metastatic versus nonmetastatic breast and prostate cancer xenografts. Neoplasia 3:143–153PubMedCrossRefGoogle Scholar
  27. 27.
    Denko NC, Fontana LA, Hudson KM, Sutphin PD, Raychaudhuri S, Altman R, Giaccia AJ 2003 Investigating hypoxic tumour physiology through gene expression patterns. Oncogene 22:5907–5914PubMedCrossRefGoogle Scholar
  28. 28.
    Bredel M, Bredel C, Juric D, Harsh GR, Vogel H, Recht LD, Sikic BI (2005) Functional network analysis reveales extended gliomagenesis pathway maps and three novel MYC-interacting genes in human gliomas. Cancer Res 65(19):8679–8689PubMedCrossRefGoogle Scholar
  29. 29.
    Tabatabai G, Bähr O, Möhle R, Eyüpoglu IY, Boehmler AM, Wischhusen J, Rieger J, Blümcke I, Weller M, Wick W (2005) Brain 128:2200–2211PubMedCrossRefGoogle Scholar
  30. 30.
    Glass R, Synowitz M, Kronenberg G, Walzlein JH, Markovic DS, Wang LP, Gast D, Kiwit J, Kempermann G, Kettenmann H (2005) Glioblastoma-induced attraction of endogenous neural precursor cells is associated with improved survival. J Neurosci 25(10):2637–2646PubMedCrossRefGoogle Scholar
  31. 31.
    Kondo T, Setoguchi T, Taga T (2004) Persistence of a small subpopulation of cancer stem-like cells in the C6 glioma cell line. Proc Natl Acad Sci USA 101(3):781–786PubMedCrossRefGoogle Scholar
  32. 32.
    Grobben B, De Deyn PP, Slegers H (2002) Rat C6 glioma as experimental model system for the study of glioblastoma growth and invasion. Cell Tissue Res 310:257–270PubMedCrossRefGoogle Scholar
  33. 33.
    Parsa AT, Chakrabarti I, Hurley PT, Chi JH, Hall JS, Kaiser MG, Bruce JN (2000) Limitations of the C6/Wistar rat intracerebral glioma model: implications for evaluating immunotherapy. Neurosurgery 47(4):993–1000PubMedCrossRefGoogle Scholar
  34. 34.
    Belién ATJ, Paganetti P, Schwab ME (1999) Membrane-type 1 matrix metalloprotease (MT1-MMP) enables invasive migration of glioma cells in central nervous system white matter. J Cell Biol 144:373–384PubMedCrossRefGoogle Scholar
  35. 35.
    Kodera T, Nakagawa T, Kubota T, Kubota M, Sato K, Kobayashi H (2000) The expression and activation of matrix metalloproteinase-2 in rat brain after implantation of C6 rat glioma cells. J Neurooncol 46:105–114PubMedCrossRefGoogle Scholar
  36. 36.
    Estève PO, Tremblay P, Houde M, St-Pierre Y, Mandeville R (1998) In vitro expression of MMP-2 and MMP-9 in glioma cells following exposure to inflammatory mediators. Biochim Biophys Acta 1403:85–96PubMedCrossRefGoogle Scholar
  37. 37.
    Senner V, Sturm A, Baur I, Schrell UHM, Distel L, Paulus W (1999) CD 24 promotes invasion of glioma cells in vivo. J Neuropathol Exp Neurol 58(8):795–802PubMedGoogle Scholar
  38. 38.
    Nishikawa R, Ji XD, Harmon RC, Lazar CS, Gill GN, Cavenee WK, Su Huang HJ (1994) A mutant epidermal growth factor receptor common in human glioma confers enhanced tumourigenicity. Proc Natl Acad Sci USA 91:7727–7731PubMedCrossRefGoogle Scholar
  39. 39.
    Vajkoczy P, Menger MD (2000) Vascular microenvironment in gliomas. J Neurooncol 50:99–108PubMedCrossRefGoogle Scholar
  40. 40.
    Pilkington GJ, Akinwunmi J, Ognjenovic N, Rogers JP (1993) Differential binding of anti-CD44 on human gliomas in vitro. Neuroreport 4:259–263PubMedCrossRefGoogle Scholar
  41. 41.
    Koshyomn S, Penar P, Wadsworth MP, Taatjes DJ (1997) Localization of CD44 at the invasive margin of glioblastomas by immunoelectron microscopy. Ultrastruct Pathol 21:517–525Google Scholar
  42. 42.
    Woodhouse EC, Amanatullah DF, Schetz JA, Liotta LA, Stracke ML, Clair T (1998) Adenosine receptor mediates motility in human melanoma cells. Biochem Biophys Res Commun 246:888–894PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2007

Authors and Affiliations

  • Markus Dehnhardt
    • 1
    • 2
  • Christoph Palm
    • 1
    • 3
  • Andrea Vieten
    • 1
  • Andreas Bauer
    • 1
  • Uwe Pietrzyk
    • 1
  1. 1.Institute of Neuroscience and Biophysics 3-Medicine, Research Centre JuelichJuelichGermany
  2. 2.Medac GmbHWedelGermany
  3. 3.Centre for Medical Image Computing (CMIC)University College LondonLondonUnited Kingdom

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