Analysis of in vitro migration of cell populations plays a key role in studying a wide range of dynamic cell behaviours. Cell migration is dependent on a multitude of signals ranging from growth factors to chemokines secreted by injured cells and/or respondent immune cells. In the last decade, several studies were addressed to investigate tropism of Mesenchymal Stromal Cells (MSC) for some types of cancers. Understanding the factors involved in regulating MSC migration towards tumours is essential to ultimately develop novel clinical strategies aiming at using MSC as vehicles to deliver antitumor drugs. However, the mechanism behind MSC migration is still in its infancy, and the main cause is the lack of quantitative methods to analyse the MSC motion. The analysis of objects motion relies on image-based tracking techniques and several methods have been recently proposed to track cells in vitro. However the vast majority of them are limited to fluorescently labelled cells. Very few tracking tools can analyze images taken using label-free microscopy techniques (e.g. phase contrast), that do not compromise the biological status of the cell and are the most common ways to observe living cells. However, to accurately track irregular-shaped, flat, and poor contrasted cells, such as MSC observed in phase-contrast, is really challenging. Furthermore, the majority of the tracking tools require relevant image processing skills, which strongly limits their practical usefulness within the biologist community. User-friendly and versatile open source software with track editing possibilities would boost live cell analysis research. In this work we introduced CellTracker, a user-friendly open-source software tool for tracking cells, including MSC. CellTracker is freely available at: http://celltracker.website/ and it works with Windows, Macintosh, and UNIX-based systems. By using CellTracker and μ-Slide Chemotaxis2D (IBIDI, Madison, Wisconsin, USA) we performed experiments to analyse the motility of MSC when seeded near to cancer cells, simply seeding MSC in a chamber communicating to another one containing conditioned medium from MG-63 osteosarcoma cells. As negative and chemotaxis controls, we used respectively fresh culture medium and culture medium supplemented with serum. From the CellTracker analysis we could not detect any significant change in MSC motility towards the culture medium with or without serum, whereas a different behaviour was observed by using culture conditioned medium taken from flasks previously containing MG-63 cells. We can therefore argue that MSC migration towards tumour cells is mediated by the presence of specific proteins (e.g. chemokines) secreted by cancer cells in the culture medium. These preliminary results pave the way for future applications such as the possibility to develop hybrid disposable diagnostic devices where biological fluids (e.g. blood) sampled from the area of interest are analysed by using the MSC motility as a cancer sensor.
Piccinini, F., Serena, D., Elisa, M., Giulio, A., Lucarelli, E., Bevilacqua, A. (2015). In vitro quantitative analysis of mesenchymal stromal cells migration towards tumours.
In vitro quantitative analysis of mesenchymal stromal cells migration towards tumours
PICCININI, FILIPPO;LUCARELLI, ENRICO;BEVILACQUA, ALESSANDRO
2015
Abstract
Analysis of in vitro migration of cell populations plays a key role in studying a wide range of dynamic cell behaviours. Cell migration is dependent on a multitude of signals ranging from growth factors to chemokines secreted by injured cells and/or respondent immune cells. In the last decade, several studies were addressed to investigate tropism of Mesenchymal Stromal Cells (MSC) for some types of cancers. Understanding the factors involved in regulating MSC migration towards tumours is essential to ultimately develop novel clinical strategies aiming at using MSC as vehicles to deliver antitumor drugs. However, the mechanism behind MSC migration is still in its infancy, and the main cause is the lack of quantitative methods to analyse the MSC motion. The analysis of objects motion relies on image-based tracking techniques and several methods have been recently proposed to track cells in vitro. However the vast majority of them are limited to fluorescently labelled cells. Very few tracking tools can analyze images taken using label-free microscopy techniques (e.g. phase contrast), that do not compromise the biological status of the cell and are the most common ways to observe living cells. However, to accurately track irregular-shaped, flat, and poor contrasted cells, such as MSC observed in phase-contrast, is really challenging. Furthermore, the majority of the tracking tools require relevant image processing skills, which strongly limits their practical usefulness within the biologist community. User-friendly and versatile open source software with track editing possibilities would boost live cell analysis research. In this work we introduced CellTracker, a user-friendly open-source software tool for tracking cells, including MSC. CellTracker is freely available at: http://celltracker.website/ and it works with Windows, Macintosh, and UNIX-based systems. By using CellTracker and μ-Slide Chemotaxis2D (IBIDI, Madison, Wisconsin, USA) we performed experiments to analyse the motility of MSC when seeded near to cancer cells, simply seeding MSC in a chamber communicating to another one containing conditioned medium from MG-63 osteosarcoma cells. As negative and chemotaxis controls, we used respectively fresh culture medium and culture medium supplemented with serum. From the CellTracker analysis we could not detect any significant change in MSC motility towards the culture medium with or without serum, whereas a different behaviour was observed by using culture conditioned medium taken from flasks previously containing MG-63 cells. We can therefore argue that MSC migration towards tumour cells is mediated by the presence of specific proteins (e.g. chemokines) secreted by cancer cells in the culture medium. These preliminary results pave the way for future applications such as the possibility to develop hybrid disposable diagnostic devices where biological fluids (e.g. blood) sampled from the area of interest are analysed by using the MSC motility as a cancer sensor.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
