Multiphoton microscopy has defined standards for 3D fluorescence and higher harmonic generation analysis of cells and tissue structures in vitro and in vivo. Compared to single-photon excited confocal microscopy, two-photon microscopy utilizes near-infrared (NIR) excitation generating twice to multi-fold enhanced tissue penetration, reduced light scattering and minimized phototoxicity and photobleaching at out-of-focus regions, yet preserves submicron spatial resolution and subcellular detail of cell and tissue structures. Using invasive tumor xenografts in the dorsal skin-fold chamber in nude mice, we here show the dynamics of tumor growth, neoangiogenesis, and tumor invasion into the adjacent tissue microenvironment. Using fluorescent labels, not only single cells but also extensively invading collective cell strands were reconstructed to move along and around preexisting blood and lymphatic vessels, not however neovessels. Using dual-color cells expressing Histone-H2B/eGFP in the nucleus and cytoplasmic RFP, the combined dynamics of collective invasion and mitotic activity defines the hallmarks of invasive growth. In vivo imaging now allows to study molecular programs controlling metastatic cancer progression are diverse in different cancers as well as within the microenvironment of a single lesion. These include amoeboid, mesenchymal and collective invasion processes as well as cellular and molecular plasticity during molecular intervention. In future studies, time-resolved two-photon microscopy will allow to gain novel insight into the mechanisms cancer progression, regression, and persistence during experimental therapy.
03 - 07 May 2008
European Society of Endocrinology