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. 1999 Aug;80(11):1697–1707. doi: 10.1038/sj.bjc.6690586

Hypoxia-induced metastasis of human melanoma cells: involvement of vascular endothelial growth factor-mediated angiogenesis

E K Rofstad 1, T Danielsen 1
PMCID: PMC2374267  PMID: 10468285

Abstract

Tumour cells exposed to hypoxia have been shown to up-regulate the expression of vascular endothelial growth factor (VEGF). The purpose of the present work was to investigate whether hypoxia-induced VEGF up-regulation can result in increased metastatic efficiency of human melanoma cells. Two melanoma lines, one showing high (A-07) and the other showing low (D-12) VEGF secretion under aerobic conditions, were included in the study. Cell cultures were exposed to hypoxia (oxygen concentrations < 10 ppm) in vitro and metastatic efficiency, i.e. lung colonization efficiency, as well as transplantability and angiogenic potential were assessed in BALB/c-nu/nu mice. Both cell lines showed significantly increased VEGF secretion under hypoxic conditions as measured by enzyme-linked immunosorbent assay. The D-12 cells showed increased metastatic efficiency, transplantability and angiogenic potential following exposure to hypoxia. The metastatic efficiency increased with the duration of the hypoxia treatment and decreased with the time after reoxygenation. The A-07 cells on the other hand showed unchanged metastatic efficiency, transplantability and angiogenic potential following exposure to hypoxia. Both cell lines showed significantly decreased metastatic efficiency and angiogenic potential in mice treated with neutralizing antibody against VEGF. These results suggest that (a) VEGF is a limiting factor for the rate of angiogenesis in low but not in high VEGF-expressing melanomas under normoxic conditions and (b) transient hypoxia might promote the development of metastases in low VEGF-expressing melanomas by upregulating the expression of VEGF and hence enhancing the angiogenic potential of the tumour cells. © 1999 Cancer Research Campaign

Keywords: angiogenesis, hypoxia, melanoma, metastasis, VEGF

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Selected References

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  1. Asano M., Yukita A., Matsumoto T., Kondo S., Suzuki H. Inhibition of tumor growth and metastasis by an immunoneutralizing monoclonal antibody to human vascular endothelial growth factor/vascular permeability factor121. Cancer Res. 1995 Nov 15;55(22):5296–5301. [PubMed] [Google Scholar]
  2. Bochner B. H., Cote R. J., Weidner N., Groshen S., Chen S. C., Skinner D. G., Nichols P. W. Angiogenesis in bladder cancer: relationship between microvessel density and tumor prognosis. J Natl Cancer Inst. 1995 Nov 1;87(21):1603–1612. doi: 10.1093/jnci/87.21.1603. [DOI] [PubMed] [Google Scholar]
  3. Brizel D. M., Scully S. P., Harrelson J. M., Layfield L. J., Bean J. M., Prosnitz L. R., Dewhirst M. W. Tumor oxygenation predicts for the likelihood of distant metastases in human soft tissue sarcoma. Cancer Res. 1996 Mar 1;56(5):941–943. [PubMed] [Google Scholar]
  4. 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]
  5. Claffey K. P., Brown L. F., del Aguila L. F., Tognazzi K., Yeo K. T., Manseau E. J., Dvorak H. F. Expression of vascular permeability factor/vascular endothelial growth factor by melanoma cells increases tumor growth, angiogenesis, and experimental metastasis. Cancer Res. 1996 Jan 1;56(1):172–181. [PubMed] [Google Scholar]
  6. Coleman C. N. Hypoxia in tumors: a paradigm for the approach to biochemical and physiologic heterogeneity. J Natl Cancer Inst. 1988 May 4;80(5):310–317. doi: 10.1093/jnci/80.5.310. [DOI] [PubMed] [Google Scholar]
  7. Dachs G. U., Stratford I. J. The molecular response of mammalian cells to hypoxia and the potential for exploitation in cancer therapy. Br J Cancer Suppl. 1996 Jul;27:S126–S132. [PMC free article] [PubMed] [Google Scholar]
  8. Denekamp J., Hobson B. Endothelial-cell proliferation in experimental tumours. Br J Cancer. 1982 Nov;46(5):711–720. doi: 10.1038/bjc.1982.263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dvorak H. F., Orenstein N. S., Carvalho A. C., Churchill W. H., Dvorak A. M., Galli S. J., Feder J., Bitzer A. M., Rypysc J., Giovinco P. Induction of a fibrin-gel investment: an early event in line 10 hepatocarcinoma growth mediated by tumor-secreted products. J Immunol. 1979 Jan;122(1):166–174. [PubMed] [Google Scholar]
  10. Dvorak H. F. Tumors: wounds that do not heal. Similarities between tumor stroma generation and wound healing. N Engl J Med. 1986 Dec 25;315(26):1650–1659. doi: 10.1056/NEJM198612253152606. [DOI] [PubMed] [Google Scholar]
  11. Feng D., Nagy J. A., Hipp J., Dvorak H. F., Dvorak A. M. Vesiculo-vacuolar organelles and the regulation of venule permeability to macromolecules by vascular permeability factor, histamine, and serotonin. J Exp Med. 1996 May 1;183(5):1981–1986. doi: 10.1084/jem.183.5.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fidler I. J., Ellis L. M. The implications of angiogenesis for the biology and therapy of cancer metastasis. Cell. 1994 Oct 21;79(2):185–188. doi: 10.1016/0092-8674(94)90187-2. [DOI] [PubMed] [Google Scholar]
  13. Folkman J. What is the evidence that tumors are angiogenesis dependent? J Natl Cancer Inst. 1990 Jan 3;82(1):4–6. doi: 10.1093/jnci/82.1.4. [DOI] [PubMed] [Google Scholar]
  14. Gasparini G., Weidner N., Maluta S., Pozza F., Boracchi P., Mezzetti M., Testolin A., Bevilacqua P. Intratumoral microvessel density and p53 protein: correlation with metastasis in head-and-neck squamous-cell carcinoma. Int J Cancer. 1993 Nov 11;55(5):739–744. doi: 10.1002/ijc.2910550507. [DOI] [PubMed] [Google Scholar]
  15. Guidi A. J., Abu-Jawdeh G., Berse B., Jackman R. W., Tognazzi K., Dvorak H. F., Brown L. F. Vascular permeability factor (vascular endothelial growth factor) expression and angiogenesis in cervical neoplasia. J Natl Cancer Inst. 1995 Aug 16;87(16):1237–1245. doi: 10.1093/jnci/87.16.1237. [DOI] [PubMed] [Google Scholar]
  16. Herlyn M. Human melanoma: development and progression. Cancer Metastasis Rev. 1990 Sep;9(2):101–112. doi: 10.1007/BF00046337. [DOI] [PubMed] [Google Scholar]
  17. Hill R. P. Tumor progression: potential role of unstable genomic changes. Cancer Metastasis Rev. 1990 Sep;9(2):137–147. doi: 10.1007/BF00046340. [DOI] [PubMed] [Google Scholar]
  18. Hlatky L., Tsionou C., Hahnfeldt P., Coleman C. N. Mammary fibroblasts may influence breast tumor angiogenesis via hypoxia-induced vascular endothelial growth factor up-regulation and protein expression. Cancer Res. 1994 Dec 1;54(23):6083–6086. [PubMed] [Google Scholar]
  19. Hockel M., Schlenger K., Aral B., Mitze M., Schaffer U., Vaupel P. Association between tumor hypoxia and malignant progression in advanced cancer of the uterine cervix. Cancer Res. 1996 Oct 1;56(19):4509–4515. [PubMed] [Google Scholar]
  20. Horsman M. R. Nicotinamide and other benzamide analogs as agents for overcoming hypoxic cell radiation resistance in tumours. A review. Acta Oncol. 1995;34(5):571–587. doi: 10.3109/02841869509094031. [DOI] [PubMed] [Google Scholar]
  21. Jang A., Hill R. P. An examination of the effects of hypoxia, acidosis, and glucose starvation on the expression of metastasis-associated genes in murine tumor cells. Clin Exp Metastasis. 1997 Sep;15(5):469–483. doi: 10.1023/a:1018470709523. [DOI] [PubMed] [Google Scholar]
  22. Kim K. J., Li B., Winer J., Armanini M., Gillett N., Phillips H. S., Ferrara N. Inhibition of vascular endothelial growth factor-induced angiogenesis suppresses tumour growth in vivo. Nature. 1993 Apr 29;362(6423):841–844. doi: 10.1038/362841a0. [DOI] [PubMed] [Google Scholar]
  23. Kreisle R. A., Ershler W. B. Investigation of tumor angiogenesis in an id mouse model: role of host-tumor interactions. J Natl Cancer Inst. 1988 Aug 3;80(11):849–854. doi: 10.1093/jnci/80.11.849. [DOI] [PubMed] [Google Scholar]
  24. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  25. Macchiarini P., Fontanini G., Hardin M. J., Squartini F., Angeletti C. A. Relation of neovascularisation to metastasis of non-small-cell lung cancer. Lancet. 1992 Jul 18;340(8812):145–146. doi: 10.1016/0140-6736(92)93217-b. [DOI] [PubMed] [Google Scholar]
  26. Mandriota S. J., Seghezzi G., Vassalli J. D., Ferrara N., Wasi S., Mazzieri R., Mignatti P., Pepper M. S. Vascular endothelial growth factor increases urokinase receptor expression in vascular endothelial cells. J Biol Chem. 1995 Apr 28;270(17):9709–9716. doi: 10.1074/jbc.270.17.9709. [DOI] [PubMed] [Google Scholar]
  27. Mattern J., Koomägi R., Volm M. Association of vascular endothelial growth factor expression with intratumoral microvessel density and tumour cell proliferation in human epidermoid lung carcinoma. Br J Cancer. 1996 Apr;73(7):931–934. doi: 10.1038/bjc.1996.166. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Melnyk O., Shuman M. A., Kim K. J. Vascular endothelial growth factor promotes tumor dissemination by a mechanism distinct from its effect on primary tumor growth. Cancer Res. 1996 Feb 15;56(4):921–924. [PubMed] [Google Scholar]
  29. Nagy J. A., Masse E. M., Herzberg K. T., Meyers M. S., Yeo K. T., Yeo T. K., Sioussat T. M., Dvorak H. F. Pathogenesis of ascites tumor growth: vascular permeability factor, vascular hyperpermeability, and ascites fluid accumulation. Cancer Res. 1995 Jan 15;55(2):360–368. [PubMed] [Google Scholar]
  30. Pötgens A. J., van Altena M. C., Lubsen N. H., Ruiter D. J., de Waal R. M. Analysis of the tumor vasculature and metastatic behavior of xenografts of human melanoma cell lines transfected with vascular permeability factor. Am J Pathol. 1996 Apr;148(4):1203–1217. [PMC free article] [PubMed] [Google Scholar]
  31. Roberts W. G., Palade G. E. Increased microvascular permeability and endothelial fenestration induced by vascular endothelial growth factor. J Cell Sci. 1995 Jun;108(Pt 6):2369–2379. doi: 10.1242/jcs.108.6.2369. [DOI] [PubMed] [Google Scholar]
  32. Rofstad E. K. Metastatic behavior of human tumors in congenitally athymic nude mice: intrinsic properties of the tumor cells and host immune reactivity. Int J Cancer. 1995 Nov 27;63(5):744–749. doi: 10.1002/ijc.2910630523. [DOI] [PubMed] [Google Scholar]
  33. Rofstad E. K. Orthotopic human melanoma xenograft model systems for studies of tumour angiogenesis, pathophysiology, treatment sensitivity and metastatic pattern. Br J Cancer. 1994 Nov;70(5):804–812. doi: 10.1038/bjc.1994.403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Rofstad E. K. Retention of cellular radiation sensitivity in cell and xenograft lines established from human melanoma surgical specimens. Cancer Res. 1992 Apr 1;52(7):1764–1769. [PubMed] [Google Scholar]
  35. Saleh M., Stacker S. A., Wilks A. F. Inhibition of growth of C6 glioma cells in vivo by expression of antisense vascular endothelial growth factor sequence. Cancer Res. 1996 Jan 15;56(2):393–401. [PubMed] [Google Scholar]
  36. Sanna K., Rofstad E. K. Hypoxia-induced resistance to doxorubicin and methotrexate in human melanoma cell lines in vitro. Int J Cancer. 1994 Jul 15;58(2):258–262. doi: 10.1002/ijc.2910580219. [DOI] [PubMed] [Google Scholar]
  37. Schwickert G., Walenta S., Sundfør K., Rofstad E. K., Mueller-Klieser W. Correlation of high lactate levels in human cervical cancer with incidence of metastasis. Cancer Res. 1995 Nov 1;55(21):4757–4759. [PubMed] [Google Scholar]
  38. Senger D. R., Galli S. J., Dvorak A. M., Perruzzi C. A., Harvey V. S., Dvorak H. F. Tumor cells secrete a vascular permeability factor that promotes accumulation of ascites fluid. Science. 1983 Feb 25;219(4587):983–985. doi: 10.1126/science.6823562. [DOI] [PubMed] [Google Scholar]
  39. Shweiki D., Itin A., Soffer D., Keshet E. Vascular endothelial growth factor induced by hypoxia may mediate hypoxia-initiated angiogenesis. Nature. 1992 Oct 29;359(6398):843–845. doi: 10.1038/359843a0. [DOI] [PubMed] [Google Scholar]
  40. Spiro I. J., Rice G. C., Durand R. E., Stickler R., Ling C. C. Cell killing, radiosensitization and cell cycle redistribution induced by chronic hypoxia. Int J Radiat Oncol Biol Phys. 1984 Aug;10(8):1275–1280. doi: 10.1016/0360-3016(84)90332-8. [DOI] [PubMed] [Google Scholar]
  41. Sundfør K., Lyng H., Rofstad E. K. Tumour hypoxia and vascular density as predictors of metastasis in squamous cell carcinoma of the uterine cervix. Br J Cancer. 1998 Sep;78(6):822–827. doi: 10.1038/bjc.1998.586. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Suzuki N., Frapart M., Grdina D. J., Meistrich M. L., Withers H. R. Cell cycle dependency of metastatic lung colony formation. Cancer Res. 1977 Oct;37(10):3690–3693. [PubMed] [Google Scholar]
  43. Takahashi Y., Kitadai Y., Bucana C. D., Cleary K. R., Ellis L. M. Expression of vascular endothelial growth factor and its receptor, KDR, correlates with vascularity, metastasis, and proliferation of human colon cancer. Cancer Res. 1995 Sep 15;55(18):3964–3968. [PubMed] [Google Scholar]
  44. Toi M., Inada K., Hoshina S., Suzuki H., Kondo S., Tominaga T. Vascular endothelial growth factor and platelet-derived endothelial cell growth factor are frequently coexpressed in highly vascularized human breast cancer. Clin Cancer Res. 1995 Sep;1(9):961–964. [PubMed] [Google Scholar]
  45. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Unemori E. N., Ferrara N., Bauer E. A., Amento E. P. Vascular endothelial growth factor induces interstitial collagenase expression in human endothelial cells. J Cell Physiol. 1992 Dec;153(3):557–562. doi: 10.1002/jcp.1041530317. [DOI] [PubMed] [Google Scholar]
  47. Vaupel P., Kallinowski F., Okunieff P. Blood flow, oxygen and nutrient supply, and metabolic microenvironment of human tumors: a review. Cancer Res. 1989 Dec 1;49(23):6449–6465. [PubMed] [Google Scholar]
  48. Walenta S., Salameh A., Lyng H., Evensen J. F., Mitze M., Rofstad E. K., Mueller-Klieser W. Correlation of high lactate levels in head and neck tumors with incidence of metastasis. Am J Pathol. 1997 Feb;150(2):409–415. [PMC free article] [PubMed] [Google Scholar]
  49. Weidner N., Semple J. P., Welch W. R., Folkman J. Tumor angiogenesis and metastasis--correlation in invasive breast carcinoma. N Engl J Med. 1991 Jan 3;324(1):1–8. doi: 10.1056/NEJM199101033240101. [DOI] [PubMed] [Google Scholar]
  50. Weidner N. Tumor angiogenesis: review of current applications in tumor prognostication. Semin Diagn Pathol. 1993 Nov;10(4):302–313. [PubMed] [Google Scholar]
  51. Yoneda J., Kuniyasu H., Crispens M. A., Price J. E., Bucana C. D., Fidler I. J. Expression of angiogenesis-related genes and progression of human ovarian carcinomas in nude mice. J Natl Cancer Inst. 1998 Mar 18;90(6):447–454. doi: 10.1093/jnci/90.6.447. [DOI] [PubMed] [Google Scholar]
  52. Young S. D., Marshall R. S., Hill R. P. Hypoxia induces DNA overreplication and enhances metastatic potential of murine tumor cells. Proc Natl Acad Sci U S A. 1988 Dec;85(24):9533–9537. doi: 10.1073/pnas.85.24.9533. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Zagzag D. Angiogenic growth factors in neural embryogenesis and neoplasia. Am J Pathol. 1995 Feb;146(2):293–309. [PMC free article] [PubMed] [Google Scholar]
  54. Zhang H. T., Craft P., Scott P. A., Ziche M., Weich H. A., Harris A. L., Bicknell R. Enhancement of tumor growth and vascular density by transfection of vascular endothelial cell growth factor into MCF-7 human breast carcinoma cells. J Natl Cancer Inst. 1995 Feb 1;87(3):213–219. doi: 10.1093/jnci/87.3.213. [DOI] [PubMed] [Google Scholar]

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