Abstract
Ethanol exposure causes neurotoxicity, where neuroinflammation has been proposed to contribute to ethanol neurotoxicity. In addition to astroglia, microglia, as resident immune cells in the central nervous system, have been implicated as a key contributor to the neuroimmune and inflammatory processes. However, little is known regarding the role of microglia in alcohol-induced neuronal dysfunction. In this chapter, we describe a method that provides an effective and unbiased global-scale analysis for relative quantitation of protein expression in microglial cells to elucidate the molecular mechanisms underlying microglial activation after ethanol exposure. The approach involves stable isotope labeling with amino acids in cell culture followed by mass spectrometric analysis of stable isotope-labeled proteins derived from cultured microglial cells and represents a powerful tool that can be used for general assessment of microglial response at the protein level.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Bates, M. E., Bowden, S. C., and Barry, D. (2002) Neurocognitive impairment associated with alcohol use disorders: implications for treatment. Exp. Clin. Psychopharm. 10, 193–212.
Nixon, K. (2006) Alcohol and adult neurogenesis: roles in neurodegeneration and recovery in chronic alcoholism. Hippocampus 16, 287–295.
Harper, C. (1998) The neuropathology of alcohol-specific brain damage, or does alcohol damage the brain? J. Neuropath. Exp. Neur. 57, 101–110.
Blanco, A. M., Valles, S. L., Pascual, M., and Guerri, C. (2005) Involvement of TLR4/type I IL-1 receptor signaling in the induction of inflammatory mediators and cell death induced by ethanol in cultured astrocytes. J. Immunol. 175, 6893–6899.
Crews, F. T., Bechara, R., Brown, L. A., Guidot, D. M., Mandrekar, P, Oak, S., et al. (2006) Cytokines and alcohol. Alcohol Clin. Exp. Res. 30, 720–730.
McDonough, K. H. (2003) Antioxidant nutrients and alcohol. Toxicology 189, 89–97.
He, J., and Crews, F. T. (2008) Increased MCP-1 and microglia in various regions of the human alcoholic brain. Exp. Neurol. 210, 349–358.
Qin, L., He, J., Hanes, R. N., Pluzarev, O., Hong, J. S., and Crews, F. T. (2008) Increased systemic and brain cytokine production and neuroinflammation by endotoxin following ethanol treatment. J. Neuroinflamm. 5, 10.
Aloisi, F. (1999) The role of microglia and astrocytes in CNS immune surveillance and immunopathology. Adv. Exp. Med. Bio. 468, 123–133.
Streit, W. J., Walter, S. A., and Pennell, N. A. (1999) Reactive microgliosis. Prog. Neurobiol. 57, 563–581.
Liu, B., and Hong, J. S. (2003) Role of microglia in inflammation-mediated neurodegenerative diseases: mechanisms and strategies for therapeutic intervention. J. Pharmacol. Exp. Ther. 304, 1–7.
Liu, B., Gao, H. M, and Hong, J. S. (2003) Parkinson’s disease and exposure to infectious agents and pesticides and the occurrence of brain injuries: role of neuroinflammation. Environ. Health Perspect. 111, 1065–1073.
Ong, S. E., Blagoev, B., Kratchmarova, I., Kristensen, D. B., Steen, H., Pandey, A., et al. (2002) Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics. Mol. Cell. Proteomics 1, 376–386.
Cheepsunthorn, P., Radov, L., Menzies, S., Reid, J. and Connor, J. R. (2001) Characterization of a novel brain-derived microglial cell line isolated from neonatal rat brain. Glia 35, 53–62.
Zhang, P., Hatter, A., and Liu, B. (2007) Manganese chloride stimulates rat microglia to release hydrogen peroxide. Toxicol. Lett. 173, 88–100.
Acknowledgments
We thank Amanda Edson and Jean Horak for technical assistance and the Florida Center of Excellence for Drug Discovery and Innovation Proteomics Facility for providing access to analytical instrumentation for proteomics analysis.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2012 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Liu, B., Barber, D.S., Stevens, S.M. (2012). Stable Isotope Labeling with Amino Acids in Cell Culture-Based Proteomic Analysis of Ethanol-Induced Protein Expression Profiles in Microglia. In: Kobeissy, F. (eds) Psychiatric Disorders. Methods in Molecular Biology, vol 829. Humana Press. https://doi.org/10.1007/978-1-61779-458-2_35
Download citation
DOI: https://doi.org/10.1007/978-1-61779-458-2_35
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-61779-457-5
Online ISBN: 978-1-61779-458-2
eBook Packages: Springer Protocols