The embryonic murine heart is a very important model of human heart development due to its four- chambered structure similar to the human heart, a short gestation period and a completely mapped genome which is easy to alter and manipulate. Lack of an appropriate imaging technology has previously hindered progress in uncovering the normal/abnormal mechanisms that govern early heart development in mice. The multi-disciplinary project described in this five-year proposal will develop, investigate and validate imaging and analysis tools to rapidly and thoroughly investigate the living embryonic murine heart at a level of spatial and temporal resolution previously not possible. These tools are based on the emerging technology of optical coherence tomography (OCT), which is capable of micrometer-scale resolution imaging of small biological tissue samples non-destructively and in real time. Compared to avian and zebrafish models, the murine embryo is far more challenging to culture in vitro in critical early looping stages and thus presents unique imaging challenges. We will develop methods and technology to culture mouse embryos that enable normal development and facilitate optical imaging. The significant potential of OCT for embryonic imaging is clear. We will develop the technology to fully realize this potential by improving imaging speed, resolution and scanning technology to enable complete characterization of the morphology and contractile and hemodynamic function of the early embryonic murine heart. Data interpretation is a key to the success of investigations using imaging. We will develop a suite of advanced tools for 2D/3D image preprocessing, analysis and visualization that will facilitate examination and comparison of the acquired image data. These tools, including noise reduction, registration, intensity inhomogeneity compensation, visualization using opacity optimization, semi-automatic and supervised 3D image segmentation, and tissue displacement and blood flow mapping, will facilitate new observations and discovery from these unique data. A set of baseline experiments will establish the usefulness of our OCT technology and image analysis tools during the stages of most dramatic cardiac morphogenesis. We will compare functional measurements obtained in vitro from OCT with those obtained in utero using micro ultrasound. We will investigate the effect of altered hemodynamics in NMHC-IIB transgenic ( and -/-) mice on heart looping. Finally, we will correlate gene expression with functional measurements made from OCT data. The end product of this effort will be a novel set of imaging tools based on OCT, partnered with improved embryo culturing technology, which have been optimized and validated for investigating the developing murine heart. The applicability of the developed technology will not only extend to other models of heart defects, but also to many other fields of developmental biology. The hypothesis tested in the validation aim will address critical open questions regarding the earliest functional changes leading to congenital heart defects.
Early medical treatments of congenital heart defects can only be developed if we understand the origins of the defects while the heart is first developing. The lack of an ideal way to image the structure and function of these tiny, almost microscopic hearts has limited our ability answer these questions. OCT imaging of embryonic mouse heart can fill this need and we will develop and validate the technology and methods to reach this potential.
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