Lockwood, NJ;
(2016)
The Global Imaging of Physiological and Pathophysiological Angiogenesis in live zebrafish using Optical Projection Tomography.
Doctoral thesis , UCL (University College London).
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Abstract
Angiogenesis is found deregulated in many pathologies, including cancer where it is required for tumour progression and metastasis. Although much has been learnt from in vitro studies of angiogenesis, it is increasingly accepted that biological processes can differ significantly between in vitro and in vivo contexts, with whole-body responses determining phenotype. I describe the development of a novel compressive sensing optical projection tomography (CS-OPT) platform using fluorescence microscopy, which enables live, whole organism imaging of adult zebrafish. Through incorporating contemporary compressive sensing algorithms the acquisition time is dramatically reduced, enabling decreased anaesthesia leading to improved zebrafish viability. The platform has been shown to provide accurate 3-dimensional quantifications of tumour volume and vascularisation using an adult zebrafish model of tumour progression. The model has a mCherry labelled vasculature and an inducible, liver specific cancer driven by the expression of oncogenic krasV12 labelled with eGFP. This model recapitulates human hepatocellular carcinoma. The platform is minimally invasive as zebrafish can be repeatedly imaged throughout development, from larvae to adult, without reducing viability. Therefore, CS-OPT should be beneficial for longitudinal mechanistic and drug development studies of tumour pro- gression and angiogenesis. With this in mind novel zebrafish models have been developed through genome editing techniques. The generation of an inducible knockout ptena zebrafish line that lacks functional ptenb was also attempted with the aim of creating a more metastatic cancer model to better reflect human disease. Furthermore, I performed studies to optimise both the anaesthetic regime and inducer treatment, which will be important for future studies. Thus, the developed CS-OPT modality is a powerful imaging platform for longitudinal mechanistic and drug development studies within whole organisms. This has been shown in the context of tumour progression and angiogenesis, but has the potential for further developmental and pathophysiological applications.
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