Skip to main content
PMC home page
British Journal of Cancer logoLink to British Journal of Cancer
. 1998;77(1):57–64. doi: 10.1038/bjc.1998.9

Multiparameter analysis of vasculature, perfusion and proliferation in human tumour xenografts.

J Bussink 1, J H Kaanders 1, P F Rijken 1, C A Martindale 1, A J van der Kogel 1
PMCID: PMC2151270  PMID: 9459146

Abstract

A method is presented in this report for concurrent analysis of vascular architecture, blood perfusion and proliferation characteristics in whole-tumour cross-sections of human larynx carcinoma and glioblastoma xenografts. Tumours were implanted subcutaneously in nude mice. After i.v. injection with Hoechst 33342 and bromodeoxyuridine (BrdUrd) as perfusion and proliferation markers, animals were killed. An antiendothelial antibody (9F1) was used to delineate vascular structures. Cross-sections were analysed by a multistep immune staining and a computer-controlled microscope scanning method. Each tumour section was stained and scanned four times (Hoechst, 9F1, BrdUrd and Fast Blue for all nuclei). When these images were combined, vasculature, perfusion and proliferation parameters were analysed. The labelling index (LI) was defined as the ratio of the BrdUrd-labelled area to the total nuclear area. The LI based on manual counting and the LI calculated by flow cytometry (FCM) were in good agreement with the LI based on surface analysis. LI decreased at increasing distance from its nearest vessel. In the vicinity of perfused vessels, the LI was 30-70% higher than near non-perfused vessels. This method shows that both vasculature/perfusion and proliferation characteristics can be measured in the same whole-tumour section in a semiautomatic way. This could be applied in clinical practice to identify combined human tumour characteristics that predict for a favourable response to treatment modifications.

Full text

PDF
58

Images in this article

59Figure 1
on p.59
59Figure 2
on p.59

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Asai A., Shibui S., Barker M., Vanderlaan M., Gray J. W., Hoshino T. Cell kinetics of rat 9L brain tumors determined by double labeling with iodo- and bromodeoxyuridine. J Neurosurg. 1990 Aug;73(2):254–258. doi: 10.3171/jns.1990.73.2.0254. [DOI] [PubMed] [Google Scholar]
  2. Begg A. C., Hofland I., Moonen L., Bartelink H., Schraub S., Bontemps P., Le Fur R., Van Den Bogaert W., Caspers R., Van Glabbeke M. The predictive value of cell kinetic measurements in a European trial of accelerated fractionation in advanced head and neck tumors: an interim report. Int J Radiat Oncol Biol Phys. 1990 Dec;19(6):1449–1453. doi: 10.1016/0360-3016(90)90357-p. [DOI] [PubMed] [Google Scholar]
  3. Begg A. C., McNally N. J., Shrieve D. C., Kärcher H. A method to measure the duration of DNA synthesis and the potential doubling time from a single sample. Cytometry. 1985 Nov;6(6):620–626. doi: 10.1002/cyto.990060618. [DOI] [PubMed] [Google Scholar]
  4. Bennett M. H., Wilson G. D., Dische S., Saunders M. I., Martindale C. A., Robinson B. M., O'Halloran A. E., Leslie M. D., Laing J. H. Tumour proliferation assessed by combined histological and flow cytometric analysis: implications for therapy in squamous cell carcinoma in the head and neck. Br J Cancer. 1992 Jun;65(6):870–878. doi: 10.1038/bjc.1992.183. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bernsen H. J., Rijken P. F., Oostendorp T., van der Kogel A. J. Vascularity and perfusion of human gliomas xenografted in the athymic nude mouse. Br J Cancer. 1995 Apr;71(4):721–726. doi: 10.1038/bjc.1995.141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brown J. M., Giaccia A. J. Tumour hypoxia: the picture has changed in the 1990s. Int J Radiat Biol. 1994 Jan;65(1):95–102. doi: 10.1080/09553009414550131. [DOI] [PubMed] [Google Scholar]
  7. GRAY L. H., CONGER A. D., EBERT M., HORNSEY S., SCOTT O. C. The concentration of oxygen dissolved in tissues at the time of irradiation as a factor in radiotherapy. Br J Radiol. 1953 Dec;26(312):638–648. doi: 10.1259/0007-1285-26-312-638. [DOI] [PubMed] [Google Scholar]
  8. Gasinska A., Wilson G. D., Urbanski K. Labelling index of gynaecological tumours assessed by bromodeoxyuridine staining in vitro using flow cytometry and histochemistry. Int J Radiat Biol. 1989 Nov;56(5):793–796. doi: 10.1080/09553008914552061. [DOI] [PubMed] [Google Scholar]
  9. Gatenby R. A., Kessler H. B., Rosenblum J. S., Coia L. R., Moldofsky P. J., Hartz W. H., Broder G. J. Oxygen distribution in squamous cell carcinoma metastases and its relationship to outcome of radiation therapy. Int J Radiat Oncol Biol Phys. 1988 May;14(5):831–838. doi: 10.1016/0360-3016(88)90002-8. [DOI] [PubMed] [Google Scholar]
  10. Haustermans K., Vanuytsel L., Geboes K., Lerut T., Van Thillo J., Leysen J., Coosemans W., van der Schueren E. In vivo cell kinetic measurements in human oesophageal cancer: what can be learned from multiple biopsies? Eur J Cancer. 1994;30A(12):1787–1791. doi: 10.1016/0959-8049(94)00252-z. [DOI] [PubMed] [Google Scholar]
  11. Hirst D. G., Denekamp J. Tumour cell proliferation in relation to the vasculature. Cell Tissue Kinet. 1979 Jan;12(1):31–42. doi: 10.1111/j.1365-2184.1979.tb00111.x. [DOI] [PubMed] [Google Scholar]
  12. Horsman M. R., Overgaard J. Overcoming tumour radiation resistance resulting from acute hypoxia. Eur J Cancer. 1992;28A(4-5):717–718. doi: 10.1016/0959-8049(92)90099-n. [DOI] [PubMed] [Google Scholar]
  13. Höckel M., Knoop C., Schlenger K., Vorndran B., Baussmann E., Mitze M., Knapstein P. G., Vaupel P. Intratumoral pO2 predicts survival in advanced cancer of the uterine cervix. Radiother Oncol. 1993 Jan;26(1):45–50. doi: 10.1016/0167-8140(93)90025-4. [DOI] [PubMed] [Google Scholar]
  14. Kaanders J. H., Pop L. A., Marres H. A., van der Maazen R. W., van der Kogel A. J., van Daal W. A. Radiotherapy with carbogen breathing and nicotinamide in head and neck cancer: feasibility and toxicity. Radiother Oncol. 1995 Dec;37(3):190–198. doi: 10.1016/0167-8140(95)01660-0. [DOI] [PubMed] [Google Scholar]
  15. Kjellen E., Joiner M. C., Collier J. M., Johns H., Rojas A. A therapeutic benefit from combining normobaric carbogen or oxygen with nicotinamide in fractionated X-ray treatments. Radiother Oncol. 1991 Oct;22(2):81–91. doi: 10.1016/0167-8140(91)90002-x. [DOI] [PubMed] [Google Scholar]
  16. Levine E. L., Renehan A., Gossiel R., Davidson S. E., Roberts S. A., Chadwick C., Wilks D. P., Potten C. S., Hendry J. H., Hunter R. D. Apoptosis, intrinsic radiosensitivity and prediction of radiotherapy response in cervical carcinoma. Radiother Oncol. 1995 Oct;37(1):1–9. doi: 10.1016/0167-8140(95)01622-n. [DOI] [PubMed] [Google Scholar]
  17. Nordsmark M., Overgaard M., Overgaard J. Pretreatment oxygenation predicts radiation response in advanced squamous cell carcinoma of the head and neck. Radiother Oncol. 1996 Oct;41(1):31–39. doi: 10.1016/s0167-8140(96)91811-3. [DOI] [PubMed] [Google Scholar]
  18. Rijken P. F., Bernsen H. J., van der Kogel A. J. Application of an image analysis system to the quantitation of tumor perfusion and vascularity in human glioma xenografts. Microvasc Res. 1995 Sep;50(2):141–153. doi: 10.1006/mvre.1995.1048. [DOI] [PubMed] [Google Scholar]
  19. Rodriguez R., Ritter M. A., Fowler J. F., Kinsella T. J. Kinetics of cell labeling and thymidine replacement after continuous infusion of halogenated pyrimidines in vivo. Int J Radiat Oncol Biol Phys. 1994 Apr 30;29(1):105–113. doi: 10.1016/0360-3016(94)90232-1. [DOI] [PubMed] [Google Scholar]
  20. Rojas A., Joiner M. C., Denekamp J. Extrapolations from laboratory and preclinical studies for the use of carbogen and nicotinamide in radiotherapy. Radiother Oncol. 1992 Jun;24(2):123–124. doi: 10.1016/0167-8140(92)90292-3. [DOI] [PubMed] [Google Scholar]
  21. Trotter M. J., Acker B. D., Chaplin D. J. Histological evidence for nonperfused vasculature in a murine tumor following hydralazine administration. Int J Radiat Oncol Biol Phys. 1989 Oct;17(4):785–789. doi: 10.1016/0360-3016(89)90067-9. [DOI] [PubMed] [Google Scholar]
  22. Webster L., Hodgkiss R. J., Wilson G. D. Simultaneous triple staining for hypoxia, proliferation, and DNA content in murine tumours. Cytometry. 1995 Dec 1;21(4):344–351. doi: 10.1002/cyto.990210406. [DOI] [PubMed] [Google Scholar]
  23. Weidner N., Carroll P. R., Flax J., Blumenfeld W., Folkman J. Tumor angiogenesis correlates with metastasis in invasive prostate carcinoma. Am J Pathol. 1993 Aug;143(2):401–409. [PMC free article] [PubMed] [Google Scholar]
  24. Wilson G. D., Dische S., Saunders M. I. Studies with bromodeoxyuridine in head and neck cancer and accelerated radiotherapy. Radiother Oncol. 1995 Sep;36(3):189–197. doi: 10.1016/0167-8140(95)01567-z. [DOI] [PubMed] [Google Scholar]
  25. Withers H. R., Taylor J. M., Maciejewski B. The hazard of accelerated tumor clonogen repopulation during radiotherapy. Acta Oncol. 1988;27(2):131–146. doi: 10.3109/02841868809090333. [DOI] [PubMed] [Google Scholar]

Articles from British Journal of Cancer are provided here courtesy of Cancer Research UK

RESOURCES