The overall goals of Core D are to provide (i) fiuorescence, confocal, and electron microscopy support, (ii) image analysis, and (iii) physiological support for lung perfusion experiments proposed in all projects. Centralization of the imaging and physiological support within a single core refiects the emphasis that P.l.s have placed on imaging and physiological studies in lung models. Core D is essential in order to fulfill the objectives of all projects. In addition to the research support. Core D personnel will also provide training for project participants in these methodologies. Core D will provide expertise, resources, and equipment for performing lung studies in the knockout mouse models and other mouse models in which proteins of interest are expressed through gene delivery via liposomes. Core D will provide expertise for the transfection of cDNAs in mouse lung microvessels using cationic liposomes. The physiological support component will provide standardized methods for quantification of lung vascular permeability in normal and genetically modified mice. This will include measurement of pulmonary capillary filtration coefficient and vessel wall albumin permeability surface-area product. The methods to be used have been developed specifically for the mouse lung. In addition, lung vascular albumin permeability and the routes of albumin transport will be assessed by electron microscopy and morphometric analysis using described methods. In terms of the imaging component. Core D will provide resources and expertise for (i) live cell and fixed specimen fluorescence, confocal, FRET (fluorescence resonance energy transfer), and TIRF (total internal reflective fluorescence) microscopy and (ii) transmission electron microscopy. Also, Core D will provide assessment of expression of transfected and liposome-delivered proteins by fluorescence and confocal microscopy of endothelial monolayers and whole mount lung sections as described.
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