Abstract
Peroxynitrite (ONOO–) and hypochlorous acid (HOCl) are two highly reactive oxygen species generated in biological systems. The overproduction of peroxynitrite or hypochlorous acid is implicated in a broad array of human pathologies including vascular, immunological, and neurodegenerative diseases. However, unambiguous detection of these reactive oxygen species has been relatively difficult due to their short biological half-lives and multiple reaction pathways. Based on their specific chemical reactions, we have developed fluorescent probes HKGreen-1 and HKOCl-1 for highly sensitive detection of peroxynitrite and hypochlorous acid, respectively. Both probes have been demonstrated to be able to discriminate corresponding reactive species from other reactive oxygen and nitrogen species (ROS and RNS) in not only chemical systems but also biological systems. The endogenous production of peroxynitrite in neuronal cells under oxygen-glucose deprivation (OGD) conditions has been visualized for the first time by utilizing HKGreen-1 probe, whilst the endogenous production of hypochlorous acid in macrophage cells upon stimulation with LPS, IFN-γ, and PMA has been imaged by utilizing HKOCl-1 probe.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Beckman, J. S., Beckman, T. W., Chen, J., Marshall, P. A. and Freeman, B. A. (1990) Apparent hydroxyl radical production by peroxynitrite: Implications for endothelial injury from nitric oxide and superoxide. Proc Natl Acad Sci USA 87, 1620–1624.
Gryglewski, R. J., Palmer, R. M. J. and Moncada, S. (1986) Superoxide anion is involved in the breakdown of endothelium-derived vascular relaxing factor. Nature 320, 454–456.
Huie, R. E. and Padmaja, S. (1993) The reaction of NO with superoxide. Free Radic Res 18, 195–199.
Pryor, W. A. and Squadrito, G. L. (1995) The chemistry of peroxynitrite: A product from the reaction of nitric oxide with superoxide. Am J Physiol Lung Cell Mol Physiol 268, 699–722.
Szabo, C., Ischiropoulos, H. and Radi, R. (2007) Peroxynitrite: Biochemistry, pathophysiology and development of therapeutics. Nat Rev Drug Discov 6, 662–680.
Pacher, P., Beckman, J. S. and Liaudet, L. (2007) Nitric oxide and peroxynitrite in health and disease. Physiol Rev 87, 315–424.
Radi, R., Peluffo, G., Alvarez, M. N., Naviliat, M. and Cayota, A. (2001) Unraveling peroxynitrite formation in biological systems. Free Radic Biol Med 30, 463–488.
Gomes, A., Fernandes, E. and Lima, J. L. F. C. (2006) Use of fluorescence probes for detection of reactive nitrogen species: A review. J Fluoresc 16, 119–139.
Wardman, P. (2007) Fluorescent and luminescent probes for measurement of oxidative and nitrosative species in cells and tissues: Progress, pitfalls, and prospects. Free Radic Biol Med 43, 995–1022.
Yang, D., Tang, Y. C., Chen, J., Wang, X. C., Bartberger, M. D., et al. (1999) Ketone-catalyzed decomposition of peroxynitrite via dioxirane intermediates. J Am Chem Soc 121, 11976–11983.
Yang, D., Wong, M. K. and Yan, Z. (2000) Regioselective intramolecular oxidation of phenols and anisoles by dioxiranes generated in situ. J Org Chem 65, 4179–4184.
Yang, D., Wang, H. L., Sun, Z. N., Chung, N. W. and Shen, J. G. (2006) A highly selective fluorescent probe for the detection and imaging of peroxynitrite in living cells. J Am Chem Soc 128, 6004–6005.
Zhou, G., Li, X., Hein, D. W., Xiang, X., Marshall, J. P., et al. (2008) Metallothionein suppresses angiotensin II-induced nicotinamide adenine dinucleotide phosphate oxidase activation, nitrosative stress, apoptosis, and pathological remodeling in the diabetic heart. J Am Coll Cardiol 52, 655–666.
Furuichi, T., Liu, W., Shi, H., Miyake, M. and Liu, K. J. (2005) Generation of hydrogen peroxide during brief oxygen-glucose deprivation induces preconditioning neuronal protection in primary cultured neurons. J Neurosci Res 79, 816–824.
Yang, D., Wang, H. L. and Shen, J. G. (2008) Unpublished Results in Dan Yang’s Research Group
Yap, Y. W., Whiteman, M. and Cheung, N. S. (2007) Chlorinative stress: An under appreciated mediator of neurodegeneration?. Cell Signal 19, 219–228.
Lapenna, D. and Cuccurullo, F. (1996) Hypochlorous acid and its pharmacological antagonism: An update picture. Gen Pharmacol 27, 1145–1147.
Steinbeck, M. J., Nesti, L. J., Sharkey, P. F. and Parvizi, J. (2007) Myeloperoxidase and chlorinated peptides in osteoarthritis: Potential biomarkers of the disease. J Orthop Res 25, 1128–1135.
Sugiyama, S., Kugiyama, K., Aikawa, M., Nakamura, S., Ogawa, H., et al. (2004) Hypochlorous acid, a macrophage product, induces endothelial apoptosis and tissue factor expression: Involvement of myeloperoxidase-mediated oxidant in plaque erosion and thrombogenesis. Arterioscler Thromb Vasc Biol 24, 1309–1314.
Sugiyama, S., Okada, Y., Sukhova, G. K., Virmani, R., Heinecke, J. W., et al. (2001) Macrophage myeloperoxidase regulation by granulocyte macrophage colony-stimulating factor in human atherosclerosis and implications in acute coronary syndromes. Am J Pathol 158, 879–891.
Hoy, A., Leininger-Muller, B., Kutter, D., Siest, G. and Visvikis, S. (2002) Growing significance of myeloperoxidase in non-infectious diseases. Clin Chem Lab Med 40, 2–8.
Kenmoku, S., Urano, Y., Kojima, H. and Nagano, T. (2007) Development of a highly specific rhodamine-based fluorescence probe for hypochlorous acid and its application to real-time imaging of phagocytosis. J Am Chem Soc 129, 7313–7318.
Shepherd, J., Hilderbrand, S. A., Waterman, P., Heinecke, J. W., Weissleder, R., et al. (2007) A fluorescent probe for the detection of myeloperoxidase activity in atherosclerosis-associated macrophages. Chem Biol 14, 1221–1231.
Sun, Z. N., Liu, F. Q., Chen, Y., Tam, P. K. H. and Yang, D. (2008) A highly specific BODIPY-based fluorescent probe for the detection of hypochlorous acid. Org Lett 10, 2171–2174.
Brewer, G. J. (1995) Serum-free B27/neurobasal medium supports differentiated growth of neurons from the striatum, substantia nigra, septum, cerebral cortex, cerebellum, and dentate gyrus. J Neurosci Res 42, 674–683.
Robinson, K. M., Janes, M. S. and Beckman, J. S. (2008) The selective detection of mitochondrial superoxide by live cell imaging. Nat Protocols 3, 941–947.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2010 Humana Press, a part of Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Yang, D. et al. (2010). Synthetic Fluorescent Probes for Imaging of Peroxynitrite and Hypochlorous Acid in Living Cells. In: Papkovsky, D. (eds) Live Cell Imaging. Methods in Molecular Biology, vol 591. Humana Press. https://doi.org/10.1007/978-1-60761-404-3_5
Download citation
DOI: https://doi.org/10.1007/978-1-60761-404-3_5
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-60761-403-6
Online ISBN: 978-1-60761-404-3
eBook Packages: Springer Protocols