intravital imaging

Intravital imaging using fluorescent proteins

Before the introduction of fluorescent proteins, in vivo imaging was limited to the study of cells that were transiently labelled with vital dyes.

Stable fluorescent labelling, achieved using vectors that express fluorescent proteins, now allows the direct imaging of single cells in vivo.

Using what is termed intravital microscopy — observation of a tumour of interest either through surgically created chronic-transparent windows or
directly through the opened skin of living animals — single cancer cells have been visualized.

High-resolution INTRAVITAL VIDEO MICROSCOPY of GFP-expressing tumour cells provides a powerful tool for directly observingsteps in the metastatic process. Individual, nondividing cells as well as micro- and macrometastases can be clearly visualized and quantified. Cellular details, such as pseudopodial projections, can be clearly seen. Farina et al. observed tumour cell motility at the singlecell level, including movement in and out of blood vessels, using GFP-expressing cells.

Condeelis et al. have used GFP imaging to view cells in time-lapse images
in a single optical section using a confocal microscope.

The polarity of tumour cells, along with their response to chemotatic cytokines, has been visualized by intravital imaging. These techniques enable a greater understanding of tumour cell migration in vivo.

Imaging GFP-labelled tumour cells in blood vessels

Tumour cell trafficking in blood vessels is an important route for metastatic spread. Visualization of this process, especially in real time, can tell us more about this pathological step.

For example, following injection of tumour cells that stably express GFP into the tail vein of mice, it is possible to visualize single tumour
cells in blood vessels.

Huang et al., using lung carcinoma cells, and Li et al., using rodent mammary tumour cells, observed the interaction between GFP-expressing tumour cells and the blood-vessel wall through the use of skin window chambers in rodents.

They observed angiogenesis occurring very early on in tumour colony formation, when as few as 60–80 tumour cells were present. Increased vasodilation and changes in blood-vessel morphology were also
observed in the surrounding tissue.

When 100 cells were present, neovascularization was induced.

Moore and colleagues have also visualized angiogenesis using a GFP-expressing gliosarcoma rodent cell line. As well as inducing angiogenesis, tumour cells also metastasize through the vasculature — this
process has also been visualized.

Al-Mehdi et al. and Wong et al. observed GFP-expressing tumour cells
initiating haematogenous metastases in subpleural microvessels in intact, perfused mouse and rat lungs.

Metastatic tumour cells attached to the endothelia of pulmonary precapillary arterioles and capillaries, although extravasation of the tumour cells was rare.

Unexpectedly, early tumour colony formation was also observed within the blood vessels.

The multiple uses of fluorescent proteins to visualize cancer in vivo.

References

 Hoffman RM. Nat Rev Cancer. 2005 Oct;5(10):796-806. PMID: 16195751