Our capacity to understand cellular function in health and disease from the macro to micro to nano and even to the atomic scale depend on the capabilities of advanced instrumentation. The NIH funded projects described in this application require increased imaging resolution and multi-spectral capabilities for their brightfield and fluorescence microscopy applications to advance to the next level. A Zeiss LSM 710 34 channel laser scanning confocal microscope is requested. The instrument will be operated as a shared resource to support the research programs of a core group of NIH funded major and secondary users in the UMKC School of Dentistry, UMKC School of Nursing and University of Kansas Medical Center. The user group has a research emphasis on mineralized tissues and musculoskeletal research. To accommodate the range of project applications, the instrument will be configured for basic confocal microscopy on fixed cell and tissue specimens, and for live cell imaging, multispectral imaging, fluorescence recovery after photobleaching (FRAP) and optical sectioning/3D imaging. One of the projects that will employ this technology is investigating the mechanisms of assembly of bone extracellular matrix (ECM), using time lapse imaging with novel fluorescent probes for various bone ECM components. These studies are providing novel insights into the dynamic nature of bone matrix assembly and the role of cell motility in the assembly process. The role of the osteocyte in regulating skeletal responses to mechanical loading is the focus of three of the participating projects. Until recently, little was known about the function of this elusive cell type, due to its inaccessibility, embedded within bone. With the recent identification of osteocyte differentiation markers and development of transgenic mice with fluorescent osteocyte lineage reporters, it is possible to image these cells within their mineralized environment to an unprecedented level. One project examines the dynamic properties of osteoblasts and osteocytes using lineage reporters combined with time lapse imaging to gain insight into the process by which osteoblasts differentiate into osteocytes. Another project examines the role of the osteocyte protein, E11/gp38, in dendrite formation and in osteocyte responses to mechanical loading. A third project examines the molecular mechanisms by which osteocytes perceive mechanical loading signals and translate them into bone formative responses. Other projects involve calcium imaging in muscle, determining the molecular mechanisms regulating phosphate homeostasis and determining the molecular pathogenesis of bone inflammatory lesions in Cherubism patients. The Zeiss LSM 710 system will advance the progress of these projects by providing an unsurpassed level of resolution that is needed for 3D imaging of osteocyte networks in mineralized tissues. The multi-spectral imaging capabilities of the LSM 710 and its enhanced sensitivity for live imaging, are essential to advance this research to the next level, and will allow for simultaneous imaging of multiple molecular probes, maximizing the information gathered from each experiment. Acquisition of this technology will advance discoveries regarding musculoskeletal health and will have implications for diseases, such as osteoporosis, osteomalacia and inherited connective tissue disorders.
This application is for a Zeiss LSM 710 34 channel laser scanning confocal microscope, to be operated as a shared resource that will support the research programs of a core group of NIH funded major users and secondary users, located in the UMKC School of Dentistry, UMKC School of Nursing and University of Kansas Medical Center. The research emphasis of the core users is on mineralized and musculoskeletal tissues and this state of the art instrumentation will enhance their ability to generate high resolution images of muscle cells and bone cells in culture and in situ within the mineralized tissue to advance the research goals of the participating projects. Acquisition of this technology will result in new discoveries regarding bone and muscle health and will have major implications for diseases of mineralized tissues, such as osteoporosis, osteomalacia and inherited connective tissue disorders.
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