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esearch objectives

Cancer biophysics

Introduction:
The van Rheenen lab investigates how healthy and tumorigenic tissues are derived from and maintained by (cancer) stem cells, how tumor cells disseminate from primary tumors and how these disseminated tumor cells grow out at distant organs. Our group develops and utilizes state-of-the-art imaging techniques to visualize the adaptive properties of the few cells (e.g. stem and/or migratory cells) within the large population of non-metastasizing and differentiated cells that may maintain the heterogeneous tumor and metastasize. We combine the latest genetic tumor models with intravital imaging (the visualization of single cells in living mice) (Movie1). For this, we have developed techniques (published in Nature Methods and highlighted by Nature and international media such as BBC, Reuters, La Recherche) to trace individual tumor cells within the primary tumor and at distant organs in a living mouse for several weeks at subcellular resolution.

Our intravital imaging studies focus on four areas of research:
1) How is healthy tissue formed and maintained by stem cells (focused on intestinal and mammary tissue)?
2) How is heterogeneity of tumors formed and maintained (e.g. imaging of (cancer) stem cells)?
3) How and why do tumor cells escape from primary tumors?
4) How and why can tumor cells form metastasis at a distant organ?

Imaging techniques:
The van Rheenen group uses (spinning disk) confocal and multi-photon microscopes to visualize (cancer) cells in vitro and in living mice. High-end imaging techniques that we commonly use include FLIM, FRET, FRAP, FLIP, live cell and intravital imaging. We developed several important imaging techniques to study cancer that are now used all over the world. For example, the algorithms and optimalization of sensitized emission to visualize FRET on confocal and two-photon systems have been incorporated into Leica microscope systems. Our group also invented a widely applicable and quantitative intravital imaging technique to study the invasive behavior of large groups of cells in various tumor microenvironments. We make use of the photoconvertable protein Dendra2 to photomark individual and/or large groups of cells, allowing us to trace them over multiple days. For this, we have invented the ‘Mammary Imaging Window’ that allows repetitive intravital imaging of mammary tumors at subcellular resolution. One of the latest achievements of our lab is the development of the ‘Abdominal Imaging Window’ that allows the repetitive intravital imaging of cellular events (including division and migration of stem cells) in abdominal organs. For example in the liver, we were able to image for the first time the outgrowth of a liver metastasis over multiple days, starting from a single tumor cell. Moreover, (cancer) stem cells and their differentiated progeny can be traced at subcellular resolution for multiple weeks in living mice through this new abdominal imaging window.

Genetic mouse models:

In the van Rheenen group, we use the latest genetic mouse models for tracing fluorescently labeled cells in living mice in order to study (tumor) development in mammary and colorectal tissue. We have generated mice which express fluorescent proteins upon Cre induction (e.g. confetti model in which cells stochastically express either CFP, GFP, YFP or RFP which is passed on to all daughter cells) specifically in colorectal or mammary stem cells for our first research focus. We have also combined these models with tumor models based on the depletion of tumor suppressor genes (e.g. APC, P53), and the overexpression of oncogenes (Wnt, ErbB2, PyMT, KRasMut) that recapitulate the benign and malignant forms of human colorectal and mammary tumors for our second to fourth research focus.


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vanRheenen IVM

Movie 1: We visualize the motility of single invasive lobular carcinoma cells (P53-/-;E-cad-/-) in a living mouse. Time series of IVM images of tumor cells (green) and type I collagen (purple) were taken with a two-photon microscope. The movie is a maximum projection of a 30um-thick Z-stack at 50-80um deep into the tumor. Scale bar represents 25 um.

 

moving colorectal tumor cells in a liver pre-micrometastasis
Movie 2: Single colorectal tumor cells that arrive in the liver grow into a pre-micrometastasis where cells are motile and predominantly adjacent to liver tissue. Pre-micrometastasis grow into micrometastasis where tumor cells are non-motile. In this movie, the movement of tumor cells in pre-micrometastasis is visible. This movie is taken through the Abodominal Imaging Window (Ritsma et al, Sci Trans Med, 2012). Scale bar is 25 um.

 

 

 

 

 

 

 

 

 

 

image description

Figure 2: intravital imaging of metastasis. Top panel: Through a mammary gland imaging window, we image tumors for multiple days. By photomarking a population of tumor cells within a tumor (here we photomarked a square, which is depicted as green), we can track the cells for multiple days (here 6 (depicted as blue) and 24 (depicted as red) hours later), and observe migration, invasion and intravasation. Lower left panel: Tumors in which the tumor cells express GFP were imaged with intravital imaging. The blood vessels were labelled with fluorescent dextrans and are here depicted in red. Lower right panel, a tumor cell that enter the blood. The periphery of the blood vessel is shown in red.