NETWORK ANALYSIS OF NITRIC OXIDE PATHWAY IN ENDOTHELIAL CELLS

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Nitric oxide (NO) plays multiple roles in biology, functioning in vascular tone regulation, host immune defense, neurotransmission, and other biological responses. NO is synthesized from arginine by three NO synthase (NOS) isoforms: neuronal, inducible and endothelial. The NO metabolic pathway in vivo is very complex. In addition to producing NO, arginine is a precursor for synthesis of urea, polyamines, creatine phosphate, glutamate and oxo-glutarate. Arginine can be transported from blood into cells by cationic amino acid transporter isoforms. Arginine is synthesized from citrulline by successive actions of argininosuccinate synthetase and argininosuccinate lyase, the third and fourth enzymes of the urea cycle. Furthermore, NO can be oxidized to nitrite, nitrate, and peroxynitrite. To infer the topology of this complex pathway and to determine the mode of action, we have adopted the approach of network perturbation expression profiling analysis at steady-state. We have assigned 37 possible metabolites (or nodes) in the network. Capillary zone electrophoresis is the technology of choice to measure simultaneously multiple metabolites in the steady-state after perturbation. We have obtained time course data after perturbing arginine, ornithine, and Ca2+ levels. We are collaborating with Drs. James J. Collins and Timothy S. Gardner in the Department of Biomedical Engineering at Boston University on data analysis. The network approach will provide us a better understanding of the regulation of NO production in vivo.