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Beneficial Effect of Astragalosides on Stroke Condition Using PC12 Cells under Oxygen Glucose Deprivation and Reperfusion

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Abstract

Astragalosides (AST) are reported to be neuroprotective in focal cerebral ischemic models in vivo. In this study, the direct effect of AST against oxygen and glucose deprivation (OGD) including neuronal injury and the underlying mechanisms in vitro were investigated. 5 h OGD followed by 24 h of reperfusion [adding back oxygen and glucose (OGD-R)] was used to induce in vitro ischemia reperfusion injury in differentiated rat pheochromocytoma PC12 cells. AST (1, 100, and 200 µg/mL) were added to the culture after 5 h of the OGD ischemic insult and was present during the reoxygenation phases. A key finding was that OGD-R decreased cell viability, increased lactate dehydrogenase, increased reactive oxygen species, apoptosis, autophagy, functional impairment of mitochondria, and endoplasmic reticulum stress in PC12 cells, all of which AST treatment significantly reduced. In addition, AST attenuated OGD-R-induced cell loss through P38 MAPK activation a neuroprotective effect blunted by SB203580, a specific inhibitor of P38 MAPK. Our data suggest that both apoptosis and autophagy are important characteristics of OGD-R-induced PC12 death and that treating PC12 cells with AST blocked OGD-R-induced apoptosis and autophagy by suppressing intracellular oxidative stress, functional impairment of mitochondria, and endoplasmic reticulum stress. Our data provide identification of AST that can concomitantly inhibit multiple cells death pathways following OGD injuries in neural cells.

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Abbreviations

AST:

Astragalosides

ATF6:

Activating transcription factor 6

ATT:

3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

DEVD-PNA:

Ac-ASP-Glu-Val-ASP-P-nitroanilide

ER:

Endoplasmic reticulum

GRP78:

Glucose-regulated protein 78 (or Bip)

GSK-3β:

Glycogen synthase kinase 3 beta

IRE1:

Insositol-requiring kinase 1

JC-1:

5,5′,6,6′-tetrachloro1,1′,3,3′-tetraethylbenzimidazolyl-carbocyanine iodide

LC3B:

Microtubule-associated protein 1 light chain 3

LDH:

Lactate dehydrogenase

LEHD-PNA:

Ac-Leu-Glu-His-ASP-P-nitroanilide

MMP:

Mitochondrial membrane permeabilization

NAC:

N-acetylcysteine

OGD-R:

Oxygen-glucose deprivation followed by reperfusion

PERK:

RNA-activated protein kinase-like ER resident kinase

RIPA:

Radioimmunoprecipitation assay

ROS:

Reactive oxygen species

RPMI:

Roswell Park Memorial Institute

Δφm:

Mitochondrial transmembrane potential

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Acknowledgments

This work was supported by Grant NSC 101-2314-B-218-001-MY3 from the Taiwan National Council of Science (Taipei) (to C. P. Chang) and the grant MFHR10115 from Chi Mei Medical Center (Tainan, Taiwan) (to B. Y. Chio).

Conflict of interest

The authors declared no conflicts of interest.

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Authors and Affiliations

  1. Department of Chinese Medicine, Chi Mei Medical Center, Tainan, 710, Taiwan, Republic of China

    Bi-Ying Chiu, Jung-Sheng Yu & Yao-Chin Hsu

  2. Department of Biotechnology, Southern Taiwan University of Science and Technology, Tainan, 710, Taiwan, Republic of China

    Ching-Ping Chang & Mao-Tsun Lin

  3. The Ph.D. Program for Neural Regenerative Medicine, Taipei Medical University, Taipei, 110, Taiwan, Republic of China

    Ching-Ping Chang & Mao-Tsun Lin

  4. School of Medicine, Taipei Medical University, Taipei, 110, Taiwan, Republic of China

    Jia-Wei Lin

  5. Neurosurgical Department, Taipei Medical University Shuang-Ho Hospital, New Taipei City, 23561, Taiwan, Republic of China

    Jia-Wei Lin

  6. Department of Medical Research, Chi Mei Medical Center, Tainan, 710, Taiwan, Republic of China

    Wen-Pin Liu & Mao-Tsun Lin

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  1. Bi-Ying Chiu
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Correspondence to Yao-Chin Hsu or Mao-Tsun Lin.

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Chiu, BY., Chang, CP., Lin, JW. et al. Beneficial Effect of Astragalosides on Stroke Condition Using PC12 Cells under Oxygen Glucose Deprivation and Reperfusion. Cell Mol Neurobiol 34, 825–837 (2014). https://doi.org/10.1007/s10571-014-0059-4

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  • DOI: https://doi.org/10.1007/s10571-014-0059-4

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