Elsevier

Brain Research

Volume 1346, 30 July 2010, Pages 224-236
Brain Research

Research Report
Neurotrophic and neurorescue effects of Echinacoside in the subacute MPTP mouse model of Parkinson's disease

https://doi.org/10.1016/j.brainres.2010.05.018Get rights and content

Abstract

Many experiments support the notion that augmentation of neurotrophic factors' (NTFs) activity, especially glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF) could prevent or halt the progress of neurodegeneration in Parkinson's disease (PD). However, application of NTFs as therapeutic agents for PD is hampered by the difficulty in delivering them to specific brain regions safely and effectively. Another potential strategy is to stimulate the endogenous expression of NTFs. In this study, we investigated the effects of Echinacoside (ECH), a monomer extracted from herbs, on rescuing dopaminergic function in 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-lesioned mice. We found that oral administration of ECH (30 mg/kg/day for 14 days) to MPTP-treated mice, commencing after impairment of the nigrstriatal system, suppressed the reduction of nigral dopaminergic neurons, striatal fibers, dopamine and dopamine transporter to 134.24%, 203.17%, 147.25% and 154.72 of MPTP-lesioned animals respectively (p < 0.05). There was a relative elevation in expression of GDNF and BDNF mRNA (2.94 and 3.75-fold) and protein (184.34% and 185.93%) in ECH treated mice compared with vehicle-treated MPTP-lesioned mice (p < 0.05). In addition, the apoptosis cells and Bax/Bcl-2 ratio of mRNA and protein in MPTP-lesioned mice significantly increased, and these effects could be prevented by ECH. At the 7th and 14th days of ECH treatment, the gait disorder displayed obvious improvement (p < 0.05). These findings demonstrate that ECH is probably a novel, orally active, non-peptide inducer of NTFs and inhibitor of apoptosis, and they provide preclinical support for therapeutic potential of this compound in the treatment of PD.

Introduction

Parkinson's disease (PD) is characterized by progressive dopaminergic neuronal cell death in the substantia nigra, resulting in severe motor deficits. In contrast to neuroprotective strategies, which aim to slow degeneration and thus must be initiated before significant damage has occurred, neurorescue or restorative therapies aim to obliterate neuronal deficits and degeneration after the initiation of impairments. Neurotrophic factors (NTFs) are secreted proteins that regulate the survival, functional maintenance and phenotypic development of neuronal cells, in particular glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF) are widely recognized as potent survival factors for the dopaminergic neurons and the nigrostriatal pathway that degenerate in PD. While the neuroprotective or neurorescue activities of these NTFs have been demonstrated in a number of in vitro and in vivo experiments (Peterson and Nutt, 2008), their application as therapeutic agents for PD has been hampered by the difficulties in delivering them into the brain because the macromolecule peptides cannot pass the brain blood barrier (BBB). Thus clinical trials testing treatment of PD using either surgery or gene therapy must solve the problem of efficient delivery and expression of NTFs (Dietz et al., 2006, Torp et al., 2006). As an alternative to such invasive approaches, a potential strategy is to stimulate the endogenous expression of NTFs. Recently, several non-peptide NTF inducers, such as XIB4035 and PYM50028, have been shown to attenuate the loss of dopaminergic neurons and might have beneficial effects in the treatment of PD (Tokugawa et al., 2003, Visanji et al., 2008). These low molecular weight agents can be administered systemically, and they would thus overcome most of the problems associated with the efficacy and safety of delivering NTFs into the brain by such methods as induction of NTF expression by viral vectors or with the use of encapsulated NTF producing cells (Bespalov and Saarma, 2007). Thus, molecules that either induce the endogenous expression of NTFs or enhance their signaling have been given increasing attention as alternative therapeutic options for PD.

Echinacoside (ECH, Fig. 1) is one of the principal constituents of phenylethanoid glycosides (PhGs) extracted from a famous traditional Chinese medicine, Herba Cistanches (the stems of Cistanche deserticola, Cistanche salsa or Cistanche tubulosa). It is also the main phenolic component in the roots of Echinacea angustifolia and E. pallida (Pellati et al., 2004), which are widely used in Europe and North America for their immunoregulation properties (Hostettmann, 2003).As a polyphenol natural product ECH has various bioactivities, such as powerful antioxidant (Sloley et al., 2001, Dalby-Brown et al., 2005), free radical scavenging and reduce the amount of NO radical generated by activated macrophages (Xiong et al., 2000) so that it can protect against hepatotoxicity (Wu et al., 2007) and acute lung injury in rats (Zhang et al., 2007, Xiong et al., 1998). It also has anti-inflammatory, antitumor, anti-aging and improving learning memory activities (reviewed by He et al., 2009). Several studies have reported ECH has anti-apoptotic and neuroprotective properties in neurodegenerative disease, for example, ECH shows significant neuroprotective effects on H2O2 injured PC-12 cells and rescues SH-SY5Y neuronal cells from TNF-α induce apoptosis through the mitochondrial pathway (Kuang et al., 2009, Deng et al., 2004). In vivo the compound has protected dopaminergic neurons in 6-hydroxydopamine (6-OHDA)-lesioned rats through attenuating the diminution of dopamine (DA) and its intermediates (Chen et al., 2007a, Chen et al., 2007b). Another study reported that ECH had neuroprotective effects and behavioral improvement in the mouse model of MPTP-induced dopaminergic neuronal damage, while further investigation revealed that ECH induced down-regulation of caspase-3 and caspase-8 activation in MPP+-induced apoptosis of cerebellar granule neurons (Geng et al., 2007). But these studies only investigated the anti-apoptotic or neuroprotective roles of ECH by the method of pre-treatment. The questions of whether ECH affects other dopaminergic neuronal prosurvival and death-related factors (such as NTFs and the Bcl-2 family) and whether it has neurorescue effects have not yet been addressed. Through reviewing the sparse literature on NTF induction by herbs or herbal compounds, we found that Rehmannia glutinosa and catalpol, two herbs with similar functions to Cistanches according to the traditional Chinese medical theory, could induce GDNF gene and protein expressions both in vitro and in vivo then attenuate MPTP-induced neuronal degeneration of nigral-striatal dopaminergic pathway (Yu et al., 2006, Xu et al., 2010). We therefore hypothesized that ECH may also have analogous efficacy. On the other hand the subacute MPTP mouse model is a quite popular regimen which involves one injection of 30 mg/kg daily for five consecutive days. This regimen causes apoptosis and depletes striatal dopamine by 40–50% in young adult C57BL/6 mice, and the dopaminergic lesion stabilizes by 21 days after MPTP administration (Tatton and Kish, 1997, Jackson-Lewis and Przedborski, 2007). So our current study chooses this model and investigates the effects of ECH on expressions of GDNF and BDNF, two of the most potent NTFs associated with dopaminergic neurons survival, and it further demonstrates the neurorescue and anti-apoptotic capabilities of this compound after subacute MPTP lesion induction in a mouse model of PD.

Section snippets

ECH improves gait dysfunctions in a subacute MPTP mouse model of PD

Shortened stride length is one of the chief characteristics of abnormal gait in PD. The results of our study demonstrate a significant decrease in forelimb and hindlimb stride lengths in the MPTP-lesioned mice treated with vehicle (saline) at the 9th day after the last administration of MPTP, i.e., the 7th day of vehicle treatment, compared with the normal control group (p = 0.023, 0.014 , Figs. 2A, B). Meanwhile, the stride lengths of mice in the high-dose ECH (30 mg/kg/day) treatment group is

Discussion

In the present study, we demonstrate for the first time that ECH, a monomer extracted from a traditional Chinese herb, has neurorescue activities in subacute MPTP mouse model of PD as well as the potential to induce the expression of NTFs at both transcription and protein levels. After 14 days of oral administration, ECH demonstrated multifunctional effects on rescuing dopaminergic neurons function, relieving the reduction of DAT, inhibiting the growth in number of apoptosis cells in SNpc and

Animal protocols

All procedures were approved by the Animal Ethical Committee of Zhongshan Hospital Fudan University and carried out in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals.

Experiments were conducted using male C57BL/6 mice purchased from the Shanghai Slac Laboratory Animal Company (Shanghai, China) at 10 weeks of age and weighing 24–26 g. The animals were maintained in standard conditions (12 ⁄ 12 h light ⁄ dark cycle, 21 ± 2 °C, and relative humidity of

Acknowledgments

This work was supported by the Scientific and Technical Supporting Program, the Ministry of Science and Technology of China (No. 2006BAI04A11-3) and the Medical Leader Sponsorship by Shanghai Municipal Government (No. 2007-057).We thank Professor Fang Huang and Professor Danian Zhu for their guidance on the experiments and manuscript.

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