Assessing tumor microenvironment of metastasis doorway-mediated vascular permeability associated with cancer cell dissemination using intravital imaging and fixed tissue analysis

George S. Karagiannis, Jessica M. Pastoriza, Lucia Borriello, Rojin Jafari, Anouchka Coste, John S. Condeelis, Maja H. Oktay, David Entenberg

Research output: Contribution to journalArticle

1 Scopus citations

Abstract

The most common cause of cancer related mortality is metastasis, a process that requires dissemination of cancer cells from the primary tumor to secondary sites. Recently, we established that cancer cell dissemination in primary breast cancer and at metastatic sites in the lung occurs only at doorways called Tumor MicroEnvironment of Metastasis (TMEM). TMEM doorway number is prognostic for distant recurrence of metastatic disease in breast cancer patients. TMEM doorways are composed of a cancer cell which over-expresses the actin regulatory protein Mena in direct contact with a perivascular, proangiogenic macrophage which expresses high levels of TIE2 and VEGF, where both of these cells are tightly bound to a blood vessel endothelial cell. Cancer cells can intravasate through TMEM doorways due to transient vascular permeability orchestrated by the joint activity of the TMEM-associated macrophage and the TMEM-associated Mena-expressing cancer cell. In this manuscript, we describe two methods for assessment of TMEM-mediated transient vascular permeability: intravital imaging and fixed tissue immunofluorescence. Although both methods have their advantages and disadvantages, combining the two may provide the most complete analyses of TMEM-mediated vascular permeability as well as microenvironmental prerequisites for TMEM function. Since the metastatic process in breast cancer, and possibly other types of cancer, involves cancer cell dissemination via TMEM doorways, it is essential to employ well established methods for the analysis of the TMEM doorway activity. The two methods described here provide a comprehensive approach to the analysis of TMEM doorway activity, either in naïve or pharmacologically treated animals, which is of paramount importance for pre-clinical trials of agents that prevent cancer cell dissemination via TMEM.

Original languageEnglish (US)
Article numbere59633
JournalJournal of Visualized Experiments
Volume2019
Issue number148
DOIs
StatePublished - Jun 2019

Keywords

  • Blood vessel permeability
  • Cancer Research
  • High molecular weight dextran
  • Immunofluorescence
  • Immunofluorescence
  • Intravital imaging
  • Issue 148
  • Metastasis
  • Tumor microenvironment of metastasis (TMEM)

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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