NETWORK ANALYSIS OF NITRIC OXIDE PATHWAY IN ENDOTHELIAL CELLS

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。一氧化氮(NO)在生物学中发挥多种作用，参与血管张力调节、宿主免疫防御、神经传递等生物反应。精氨酸通过神经型、诱导型和内皮型三种一氧化氮合酶亚型合成NO。NO在体内的代谢途径非常复杂。除了产生一氧化氮，精氨酸还是合成尿素、多胺、磷酸肌酸、谷氨酸和氧化戊二酸的前体。精氨酸可以通过阳离子氨基酸转运体异构体从血液中转运到细胞中。精氨酸是由瓜氨酸通过精氨酸琥珀酸合成酶和精氨酸琥珀酸裂解酶的连续作用而合成的，精氨酸琥珀酸裂解酶是尿素循环的第三和第四酶。此外，NO可以被氧化成亚硝酸盐、硝酸盐和过氧亚硝酸盐。为了推断这一复杂通路的拓扑结构和确定作用模式，我们采用了稳态网络扰动表达谱分析的方法。我们已经在网络中指定了37种可能的代谢物(或节点)。毛细管区带电泳法是在扰动后的稳态下同时测定多种代谢物的首选技术。在对精氨酸、鸟氨酸和钙离子水平进行扰动后，我们得到了时程数据。我们正在与波士顿大学生物医学工程系的詹姆斯·J·柯林斯博士和蒂莫西·S·加德纳博士合作进行数据分析。网络方法将使我们更好地了解体内NO产生的调节。

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.