The broad goal of this project is to define the molecular anatomy of the vasculature in multiple organs and to provide distinct molecular signatures unique to each vasculature. Considerable molecular diversity exists at the microvascular endothelial cell surface among organs, including the expression of organ-specific proteins which can provide key basic data necessary to investigate the mechanisms underlying the unique expression patterns as well as organ-specific vascular targets that can be used for site-specific drug delivery and molecular imaging. Defining the vascular proteome, discovering organ-specific targets, and developing suitable organ-specific probes are the goals of this project.
The specific aims of this project are: i) To combine novel subfractionation and mass spectrometry techniques to map the accessible vascular proteome of normal major organs and, through comparative analysis, uncover tissue-modulated and possibly organ-induced vascular proteins;ii) To generate and characterize antibodies specific for organ-induced vascular proteins as a means of determining the degree of molecular diversity of endothelia, performing detailed expression profiling, and creating potential tissue-targeting probes;and iii) To validate the utility of any apparent organ-specific targets and their antibodies by assessing organ-specific delivery and endothelial cell processing in vivo. In this proposal, we combine hypothesis-driven investigation with global analytical tools to establish better the validity of the hypothesis of tissue-modulated endothelial cell diversity in vivo. We use in silico subtractive and bioinformatic filters to reduce data complexity by orders of magnitude to focus on a manageable subset of proteins appearing to be both tissue-modulated and IV-accessible. Target-specific antibodies will validate IV-accessible endothelial cell surface targets via tissue expression profiling and whole body SPECT imaging in vivo. Intravital microscopy will be used to visualize endothelial cell processing of targeting antibodies in vivo. Such basic, molecular information is critical for establishing vascular function and is required for research in vascular biology, pharmacology, physiology, and development. This project is likely to provide clinically important targets potentially useful for organ-specific drug and gene delivery in vivo.
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