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Effects of Obovatol on GSH Depleted Glia-Mediated Neurotoxicity and Oxidative Damage

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An Erratum to this article was published on 11 August 2011

Abstract

Earlier studies indicate that obovatol (OBO), isolated from a medicinal herb Magnolia obovata, has anti-inflammatory and anti-oxidative properties. Depletion of glutathione (GSH) in glial cells with the γ-glutamylcysteine synthase inhibitor D,L-buthionine-S,R-sulfoximine (BSO) is known to produce oxidative stress which, in turn, induces these cells to secrete inflammatory cytokines and other neurotoxic substances. In the present study, we investigated the ability of OBO to protect SH-SY5Y neuroblastoma cells from this effect. Human microglia, astrocytes and their surrogate THP-1 and U373 cell lines were activated by treatment with BSO. Such treatment depleted their intracellular GSH and increased levels of damage to DNA, lipids and proteins (8-OHdG, lipid peroxide, protein carbonyls and 3-nitrotyrosine), and activated the inflammatory pathways P38 MAP kinase and NFκB. These are accompanied by release of proinflammatory factors such as TNFα, IL-6 and nitric oxide. Their conditioned media were toxic to SH-SY5Y cells. All these effects were attenuated by pre-treatment with OBO. Prior treatment of SH-SY5Y cells with OBO also attenuated THP-1 or U373 conditioned media neurotoxicity and also reduced oxidative damage produced by treatment with hydrogen peroxide or BSO. Prior treatment with OBO potentiated survival of SH-SY5Y cells exposed to conditioned medium from BSO-treated THP-1, U373 cells, microglia and astrocytes. The data indicate that OBO could be anti-inflammatory, anti-oxidative and neuroprotective, and be an effective agent for inhibiting pathogenesis in neurological diseases such as Alzheimer disease, Parkinson disease and amyotrophic lateral sclerosis in which glial-mediated neuroinflammation and oxidative stress are thought to contribute to disease progression.

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Acknowledgments

This research was supported by the Pacific Alzheimer Research Foundation.

Conflict of interest

None.

Author information

Authors and Affiliations

  1. Kinsmen Laboratory of Neurological Research, University of British Columbia, 2255 Wesbrook Mall, Vancouver, British Columbia, Canada, V6T 1Z3

    Moonhee Lee, Edith McGeer & Patrick L. McGeer

  2. Laboratory of Chemical Biology and Genomics, Korea Research Institute of Bioscience and Biotechnology, University of Science and Technology, Daejeon, Korea

    Byoung-Mok Kwon

  3. Department of Pharmacology, Kyungpook National University School of Medicine, Brain Science and Engineering Institute, Daegu, Korea

    Kyoungho Suk

Authors
  1. Moonhee Lee
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  2. Byoung-Mok Kwon
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  3. Kyoungho Suk
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  4. Edith McGeer
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  5. Patrick L. McGeer
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Corresponding author

Correspondence to Patrick L. McGeer.

Additional information

This research was supported by the Pacific Alzheimer Research Foundation.

An erratum to this article can be found at http://dx.doi.org/10.1007/s11481-011-9305-4.

Electronic supplementary materials

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Supplementary Figure 1

Effects of treatment with 30 μM obovatol (OBO) or 500 μM BSO on viability changes of THP-1 cells (A), U373 cells (B), microglia (C) and astrocytes (D). Four types of cells were directly treated with OBO at 30 μM or BSO at 500 μM for 2 days and MTT assay was employed to examined their viability. Values are mean±SEM, n = 4. One-way ANOVAs were carried out to test for significance. Note that there were no viability change in all cells in the presence of OBO or BSO for 2 days. (DOC 114 kb)

Supplementary Figure 2

Effects of pretreatment with 30 μM obovatol (OBO) on viability changes of undifferentiated or differentiated SH-SY5Y cells induced by conditioned medium from BSO-stimulated THP-1 cells (A), U373 cells (B), microglia (C) and astrocytes (D). THP-1 cells, U373 cells, microglia or astrocytes were pre-treated with OBO at 30 μM for 2 h and they were subsequently exposed to BSO at 500 μM for 2 days. Their conditioned medium was transferred to undifferentiated or differentiated SH-SY5Y cells with 5 μM retinoic acid for 4 days. MTT assay was used to examined their viability. Values are mean±SEM, n = 4. One-way ANOVAs were carried out to test for significance. Note that there were no viability differences between undifferentiated and differentiated SH-SY5Y cells in the same condition. (DOC 136 kb)

Supplementary Figure 3

Effects of filtration of BSO-stimulated THP-1 or U373 conditioned medium on viability changes of SH-SY5Y cells. After THP-1 cells (A) or U373 cells (B) were stimulated with 500 μM BSO for 2 days their conditioned medium was passed through a 3KDa MW cut-off filters. The filtered medium was transferred to SH-SY5Y cells and MTT assay was used to examine their viability in 3 days. Values are mean±SEM, n = 4. One-way ANOVAs were carried out to test for significance. * P < 0.01 for BSO-stimulated group compared with the control (CON) group and ** P < 0.01 for BSO + 3KDa group compared with the BSO group. Note that there was attenuation in 3KDa MW filter group compared with the unfiltered group; 25% in THP-1 cells and 15% in U373 cells. (DOC 77 kb)

Supplementary Figure 4

Effects of 30 μM obovatol (OBO) on levels of TNFμ or nitrite ions released from BSO-stimulated U373 cells (A,B) or astrocytes (C,D). After stimulation with BSO in the presence or absence of prior treatment with OBO for 2 h, the cell medium was collected to measure levels of TNFμ (A,C) and nitrite (B,D). Values are mean±SEM, n = 4. One-way ANOVAs were carried out to test for significance. ** P < 0.01 for OBO group compared with the stimulated (BSO) group. (DOC 110 kb)

Supplementary Figure 5

Effects of treatment with various concentration of H2O2 for 3 days on the viability of SH-SY5Y cells. For experimental protocols please see materials and methods section or main text. Values are mean±SEM, n = 4. A one-way ANOVA was carried out to test for significance. ** P < 0.01 for 3–300 μM treatment group compared with CON group. (DOC 54 kb)

Supplementary Figure 6

Concentration of H2O2 12 h after reaction with 30 μM obovatol (OBO) with 5 μM H2O2. OBO was incubated with H2O2 in PBS for 12 h at 37°C and the concentration of H2O2 was measured as described in the materials and methods section. White bars: 0 h and black bars: 12 h incubation. Values are mean±SEM, n = 4. A one-way ANOVA was carried out to test for significance. ** P < 0.01 compared with the same group at 0 h. (DOC 48 kb)

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Lee, M., Kwon, BM., Suk, K. et al. Effects of Obovatol on GSH Depleted Glia-Mediated Neurotoxicity and Oxidative Damage. J Neuroimmune Pharmacol 7, 173–186 (2012). https://doi.org/10.1007/s11481-011-9300-9

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