Share This Article:

Over-Expressing Prohibitin (PHB) in Neuronal Cultures Exacerbates Cell Death Following Hydrogen Peroxide and L-Glutamic Acid Induced Injury

Abstract Full-Text HTML XML Download Download as PDF (Size:2707KB) PP. 149-160
DOI: 10.4236/nm.2014.54018    2,275 Downloads   2,670 Views   Citations


Using proteomics, previous work in our laboratory identified five mitochondrial related proteins [citrate synthase (CS), glucose-regulated protein 75 (GRP75), heat shock protein 60 (HSP60), prohibitin (PHB), voltage-dependent anion channel 1 (VDAC1)] to be differentially expressed in primary cortical neuronal cultures following preconditioning treatments [1] [2]. To investigate a protective or damaging role of these five proteins in neurons, we used RNAi constructs to knockdown and adenoviral vectors to over-express the proteins in cortical neuronal cultures prior to exposure to three ischemia-related injury models: excitotoxicity (L-glutamic acid), oxidative stress (hydrogen peroxide) and in vitro ischemia (oxygen-glucose deprivation). We observed that down-regulating these mitochondrial proteins had no effect on neuronal viability, in any injury model. By contrast, over-expression of PHB exacerbated cell death in the hydrogen peroxide and L-glutamic acid injury models. These findings indicate that PHB plays a neurodamaging role following oxidative and excitotoxic stress and suggests that the protein is a potential therapeutic target for the design of drugs to limit neuronal death following cerebral ischemia and other forms of brain injury.

Conflicts of Interest

The authors declare no conflicts of interest.

Cite this paper

Teoh, J. , Boulos, S. , Chieng, J. , Knuckey, N. and Meloni, B. (2014) Over-Expressing Prohibitin (PHB) in Neuronal Cultures Exacerbates Cell Death Following Hydrogen Peroxide and L-Glutamic Acid Induced Injury. Neuroscience and Medicine, 5, 149-160. doi: 10.4236/nm.2014.54018.


[1] [1] Meloni, B.P., Tilbrook, P.A., Boulos, S., Arthur, P.G. and Knuckey, N.W. (2006) Erythropoietin Preconditioning in Neuronal Cultures: Signaling, Protection from in Vitro Ischemia, and Proteomic Analysis. Journal of Neuroscience Research, 83, 584-593.
[2] Meloni, B.P., Van Dyk, D., Cole, R. and Knuckey, N.W. (2005) Proteome Analysis of Cortical Neuronal Cultures Following Cycloheximide, Heat Stress and MK801 Preconditioning. Proteomics, 5, 4743-4753.
[3] O’Donnell, M.J., et al. (2010) Risk Factors for Ischaemic and Intracerebral Haemorrhagic Stroke in 22 Countries (the INTERSTROKE Study): A Case-Control Study. Lancet, 376, 112-123.
[4] Schretzman, D. (2001) Acute Ischemic Stroke. Dimensions of Critical Care Nursing, 20, 14-21.
[5] Maragos, W.F. and Korde, A.S. (2004) Mitochondrial Uncopling as a Potential Therapeutic Target in Acute Central Nervous System Injury. Journal of Neurochemistry, 91, 257-262.
[6] Galluzzi, L., Morselli, E., Kepp, O. and Kroemer, G. (2009) Targeting Post-Mitochondrial Effectors of Apoptosis for Neuroprotection. Biochimico et Biophysica Acta, 1787, 402-413.
[7] Meloni, B.P., Majda, B.T. and Knuckey, N.W. (2001) Establishment of Neuronal in Vitro Models of Ischemia in 96-Well Microtiter Strip-Plates That Result in Acute, Progressive and Delayed Neuronal Death. Neuroscience, 108, 17-26.
[8] He, T.C., et al. (1998) A Simplified System for Generating Recombinant Adenoviruses. Proceedings of the National Academy of Sciences USA, 95, 2509-2514.
[9] Boulos, S., Meloni, B.P., Arthur, P.G., Bojarski, C. and Knuckey, N.W. (2006) Assessment of CMV, RSV and SYN1 Promoters and the Woodchuck Post-Transcriptional Regulatory Element in Adenovirus Vectors for Transgene Expression in Cortical Neuronal Cultures. Brain Research, 1102, 27-38.
[10] Zeng, M., et al. (2001) AdEasy System Made Easier by Selecting the Viral Backbone Plasmid Preceding Homologous Recombination. Biotechniques, 31, 260-262.
[11] Luo, J., et al. (2007) A Protocol for Rapid Generation of Recombinant Adenoviruses Using the AdEasy System. Nature Protocols, 2, 1236-1247.
[12] Zhou, P., et al. (2012) Prohibitin Reduces Mitochondrial Free Radical Production and Protects Brain Cells from Different Injury Modalities. Journal of Neuroscience, 32, 583-592.
[13] Perez Velazquez, J.L., Kokarovtseva, L., Weisspapir, M. and Frantseva, M.V. (2003) Anti-Porin Antibodies Prevent Excitotoxic and Ischemic Damage to Brain Tissue. Journal of Neurotrauma, 20, 633-647.
[14] Fusaro, G., Dasgupta, P., Rastogi, S., Joshi, B. and Chellappan, S. (2003) Prohibitin Induces the Transcriptional Activity of p53 and Is Exported from the Nucleus upon Apoptotic Signaling. The Journal of Biological Chemistry, 278, 47853-47861.
[15] Terashima, M., et al. (1994) The IgM Antigen Receptor of B Lymphocytes Is Associated with Prohibitin and a Prohibitin-Related Protein. The EMBO Journal, 13, 3782-3792.
[16] Chen, X.J. and Butow, R.A. (2005) The Organization and Inheritance of the Mitochondrial Genome. Nature Reviews Genetics, 6, 815-825.
[17] Hoppins, S., Lackner, L. and Nunnari, J. (2007) The Machines That Divide and Fuse Mitochondria. Annual Review of Biochemistry, 76, 751-780.
[18] Muraguchi, T., Kawawa, A. and Kubota, S. (2010) Prohibitin Protects against Hypoxia-Induced H9c2 Cardiomyocyte Cell Death. Biomedical Research, 31, 113-122.
[19] Liu, X., et al. (2009) Prohibitin Protects against Oxidative Stress-Induced Cell Injury in Cultured Neonatal Cardiomyocyte. Cell Stress and Chaperones, 14, 311-319.
[20] Tsutsumi, T., et al. (2009) Proteomics Analysis of Mitochondrial Proteins Reveals Overexpression of a Mitochondrial Protein Chaperon, Prohibitin, in Cells Expressing Hepatitis C Virus Core Protein. Hepatology, 50, 378-386.
[21] Schleicher, M., et al. (2008) Prohibitin-1 Maintains the Angiogenic Capacity of Endothelial Cells by Regulating Mitochondrial Function and Senescence. Journal of Cell Biology, 180, 101-112.
[22] Liu, Y.H., Peck, K. and Lin, J.Y. (2012) Involvement of Prohibitin Upregulation in Abrin-Triggered Apoptosis. Evidence-Based Complementary and Alternative Medicine, 2012, Article ID: 605154.
[23] Zhang, L., Ji, Q., Ni, Z.H. and Sun, J. (2012) Prohibitin Induces Apoptosis in BGC823 Gastric Cancer Cells through the Mitochondrial Pathway. Asian Pacific Journal of Cancer Prevention, 13, 3803-3807.
[24] Voloboueva, L.A., et al. (2008) Overexpression of Mitochondrial Hsp70/Hsp75 Protects Astrocytes against Ischemic Injury in Vitro. Journal of Cerebral Blood Flow & Metabolism, 28, 1009-1016.
[25] Xu, L., Voloboueva, L.A., Ouyang, Y., Emery, J.F. and Giffard, R.G. (2009) Overexpression of Mitochondrial Hsp70/Hsp75 in Rat Brain Protects Mitochondria, Reduces Oxidative Stress, and Protects from Focal Ischemia. Journal of Cerebral Blood Flow & Metabolism, 29, 365-374.
[26] Hwang, I.K., et al. (2007) Changes in Immunoreactivity of HSP60 and Its Neuroprotective Effects in the Gerbil Hippocampal CA1 Region Induced by Transient Ischemia. Experimental Neurology, 208, 247-256.

comments powered by Disqus

Copyright © 2019 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.