Application Of Multiphoton steady state and lifetime imaging to mapping of tumour vascular architecture in vivo

Recent interest in vascular targeting and anti-angiogenic drug treatments for cancer has stimulated fundamental research regarding the modes of action of these drugs as well as studies of the development and re-modelling of the vascular network following treatment. Multiphoton fluorescence microscopy is employed for in vivo mapping of three-dimensional blood vessel distribution in tumours grown in rodent dorsal skin-flap window chamber preparations. Accurate visualisation of the vasculature in three-dimensions allows us to perform dynamic experiments in thick biological specimens in vivo. Examples of in vivo imaging of tumour vasculature are given and compared to normal tissue vasculature. The dynamic responses of blood vessels to treatment with the vascular targeting drug combretastatin A4-P are presented and discussed. The implementation of time-domain imaging by reversed stop-start time-correlated single photon counting (RSS-TCSPC) is discussed as a method for feature extraction in the presence of exogenous and endogenous fluorophores. In particular, the segmentation of the vascular network is demonstrated. Additional contrast, indicative of probe environmental factors, may also be realised. We present examples of in vivo lifetime imaging as a method to elucidate the physiological processes of the tumour microenvironment.