Intravital microscopy of tumour cells and their microenvironment through imaging windows


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Jacco van Rheenen
Hubrecht Institute, Utrecht, The Netherlands

Abstract

Complications due to metastasis are the primary reason why people die from cancer. Although histological techniques have provided important information on metastasis, they only give a static image of tumour cells and their microenvironment and thus compromise interpretation of this dynamic process. To study this dynamic process, we visualise the behaviour of single metastasising cells at subcellular resolution with two-photon intravital imaging (IVM). We have developed a Mammary Imaging Window (MIW) to image primary mammary tumours over multiple days. Through the MIW, we photomark hundreds of cells in the primary tumour that express the photoswitchable Dendra2, by switching the colour of the tumour cells from green to red. This allows us to track tumour cell migration during subsequent imaging sessions, and in addition retrace the same cells in immunohistochemical slides. Using this technique we showed that the migratory behaviour of mammary tumour cells of invasive lobular carcinomas correlates with the presence of T cells in the microenvironment.

In order to study how tumour cells arrive, survive and grow at secondary sites, we developed a new imaging window to image abdominal organs such as the liver, which is one of the primary organs for metastasis formation. Using this abdominal imaging window, we are able to visualize how individual tumour cells that arrive at the liver grow into metastases. We observe that single extravasated tumour cells proliferate and form ‘pre-micrometastases' in which cells are migratory and lack contact to neighbouring tumour cells. Subsequently, the clones condense into micrometastases in which cell migration is strongly diminished, but proliferation continues. By suppressing tumour cell migration in pre-micrometastases genetically or by drugs we reduce the number of metastases, and therefore we conclude that the migration of cells within pre-micrometastases is a novel contributing step in the formation of liver metastasis.

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