Elsevier

European Journal of Pharmacology

Volume 451, Issue 2, 13 September 2002, Pages 119-124
European Journal of Pharmacology

Protective effect of verbascoside on 1-methyl-4-phenylpyridinium ion-induced neurotoxicity in PC12 cells

https://doi.org/10.1016/S0014-2999(02)02240-9Get rights and content

Abstract

The neuroprotective effects of verbascoside, one of phenylpropanoid glucoside isolated from the Chinese herbal medicine Buddleja officinalis Maxim, on 1-methyl-4-phenylpyridinium ion (MPP+) induced apotosis and oxidative stress in PC12 neuronal cells were investigated. Treatment of PC12 cells with MPP+ for 48 h induced apoptotic death as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry, the activation of caspase-3 measured by the caspase-3 activity assay kit, the reduction in mitochondrial membrane potential with laser scanning confocal microscopy and the increase in the extracellular hydrogen peroxide level. Simultaneous treatment with verbascoside markedly attenuated MPP+-induced apoptotic death, increased extracellular hydrogen peroxide level, the activation of caspase-3 and the collapse of mitochondrial membrane potential. These results strongly indicate that verbascoside may provide a useful therapeutic strategy for the treatment of oxidative stress-induced neurodegenerative disease such as Parkinson's disease.

Introduction

The progressive neurodegeneration of nigrostriatal dopaminergic pathway is one of the principal factors accounting for the appearance of motor dysfunctions in Parkinson's disease. Even though the cause of Parkinson's disease remains answered, several lines of evidence strongly suggest involvement of oxidative stress, finally leading to neuronal death by the excessive generation of free radicals Coyle and Puttfarcken, 1993, Gotz et al., 1994. Recently, attention has been focused on a wide array of natural antioxidants that are able to scavenge free radicals and protect cells from oxidative damage, such as verbascoside, a phenylpropanoid glucoside distributed in many medicinal plants (Deepak et al., 1999). Previous studies had found many phenylpropanoid glucosides including verbascoside showed a strong free radical scavenging activity Li et al., 1999, Gao et al., 1999, Wang et al., 1996. In the present study, we have examined the possible inhibitory effect of verbascoside on neurotoxicity induced by 1-methyl-4-phenylpyridinium ion (MPP+) in cultured PC12 cells that retain dopaminergic characteristics and have been widely used for neurobiological and neurochemical studies.

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Materials

Verbascoside (Fig. 1) from Buddleja officinalis Maxim was kindly supplied by Dr. Mingju Liu (Department of Orthopaedics and Traumatology, The Chinese University of Hong Kong, Hong Kong, China). The purity of the compounds was more than 98% by high-performance liquid chromatography (HPLC) analysis. Dulbecco's modified Eagle's medium (DMEM), horse serum and fetal calf serum were purchased from Gibco BRL; poly-l-lysine, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), MPP+,

Effect of verbascoside on MPP+-induced cell death

PC12 cells were exposed to MPP+ and cell viability was assessed by the MTT reduction assay. MPP+ 200 μM killed 40–50% of the cells upon 48 h treatment and its cytotoxic effects were attenuated in the presence of verbascoside (0.1, 1 or 10 μg ml−1) (Fig. 2). Verbascoside at these concentrations exhibited cytoprotective effects in a dose-dependent manner (35%, 46% and 59% of neuroprotection, respectively) and the compound alone did not cause any apparent cytotoxicity (data not shown).

Effect of verbascoside on MPP+-induced accumulation of extracellular hydrogen peroxide

Upon

Discussion

In our study, we first demonstrate that verbascoside, isolated from B. officinalis Maxim, has significant neuroprotective effects against MPP+-induced apoptotic or necrotic death via mitochondria dysfunction in PC12 cells. The compound protected neuronal viability against MPP+-induced toxicity in a dose-dependent manner. Further, we demonstrated that verbascoside also protected against the MPP+-induced apoptosis/necrosis measured using flow cytometric analysis and caspase-3 activity assay.

Acknowledgements

This study was supported by the Doctor Fund of the Ministry of Education in China (20010001060).

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