We describe a novel approach to study tumour progression and vasculature development in vivo via global 3-D fluorescence imaging of live adult zebrafish utilising angularly multiplexed optical projection tomography with compressive sensing (CS-OPT). This method bridges a “mesocopic” imaging gap between established high resolution 3-D fluorescence microscopy techniques with whole body planar imaging and diffuse tomography.
Our new approach has been developed for imaging at multiple excitation wavelengths of non-pigmented adult zebrafish fish up to 3 cm in length. Using unfocussed illumination with two multiplexed imaging channels and iterative reconstruction of sparsely sampled OPT data, our approach requires only ~100 seconds per spectral channel for image acquisition. Using CS-OPT we imaged an inducible genetic model of liver cancer in adult transparent and fluorescent vasculature expressing zebrafish (TraNac Tg (KDR:mCherry:Fabp10-rtTA:TRE-eGFPKRASV12) ). In this disease model, addition of a chemical inducer (doxycycline) drives expression of eGFP tagged oncogenic KRASV12 in the liver of immune competent animals. We show that our novel OPT methodology enables non-invasive quantitative imaging of the development of tumour and vasculature throughout the progression of the disease and validate our results against established methods of pathology including immunohistochemistry. We have validated this new imaging approach against established methods of pathology including immunohistochemistry and further demonstrated the potential of CS-OPT through a longitudinal study of vascular development in the same zebrafish from early embryo to adulthood.
To increase the metastatic potential of our cancer model we are employing novel approaches using gene editing techniques including TALENs and CRISPRs in an attempt to produce inducible knockouts in adult zebrafish, which are currently unavailable.
We believe this imaging modality with its associated analysis and data management tools constitute a new platform for in vivo cancer studies and drug discovery in zebrafish disease models.