Intravital imaging of metastasis in zebrafish

Abstract

Metastasis is the cause of the overwhelming majority of cancer deaths. However, it remains a poorly understood process. The events at the metastatic site are especially poorly comprehended. These events are dynamic and so require intravital imaging to investigate. However, the intravital imaging of these events in mice is challenging. Sites of metastasis are often in vital organs that are inaccessible to microscopy without surgical intervention. Furthermore, circulating tumor cells are rare and are involved in many transient interactions adding to the challenge. The development of a line of zebrafish, Casper, that is transparent throughout its life suggested that zebrafish might be a powerful system for intravital imaging. I first developed novel injection and imaging techniques to study metastasis through intravital imaging in adult zebrafish. I then followed individual ZMEL1 zebrafish melanoma cells at the metastatic site over the course of two weeks as they grew from single disseminated tumor cells into macroscopic metastases. From these studies, I characterized the steps of metastasis at the metastatic site for this cell line. I also utilized transparent zebrafish embryos to uncover a new role for the oncogene YAP during metastasis. I observed that the over-expression of a Hippo-insensitive mutant of YAP (YAP-AA) promoted brain metastasis following intravenous in zebrafish embryos. I determined that YAP-AA was promoting tumor cell dispersal throughout the embryo by allowing tumor cells to escape the first capillary bed they encounter. Following intravenous injection, control cells lodge in blood vessels in the tail and cease their travel through circulation. However, YAP-AA cells are able to move through these vessels, re-enter circulation and travel to other organs, such as the brain. These observations represent a new mechanism by which tumor cells can increase their dissemination throughout an animal.