Structural characterization and neuroprotective effect of a polysaccharide from Corydalis yanhusuo

Abstract

In the present study, we investigated the neuroprotective effects of a purified Corydalis yanhusuo polysaccharide (CYP) on Aβ _(25–35)-induced neurotoxicity and explored its underlying molecular mechanisms in PC12 cells. The results showed pretreatment with CYP (25, 50, and 100 μg/ml) prior to Aβ _(25–35) exposure significantly protected PC12 cells from Aβ _(25–35)-induced cell death, lactate dehydrogenase (LDH) release, DNA fragmentation, mitochondrial dysfunction and mitochondrial cytochrome c release. Moreover, Aβ _(25–35)-induced increase of ratio between Bax and Bcl-2 protein expression was dramatically reversed by CYP pretreatment. Furthermore, the addition of CYP led to a significant repressing effect on the elevated protein expression of cleaved caspase-8, caspase-9, and caspase-3 in Aβ _(25–35)-treated PC12 cells. Taken together, these findings indicated that protective effect of CYP against Aβ _(25–35)-induced toxicity in PC12 cells was probably mediated by inhibition of apoptosis via both mitochondrial apoptotic pathway and death receptor pathway.

Introduction

Alzheimer's disease (AD) is the most widespread neurodegenerative disorders of the brain, characterized by irreversible loss of cognitive function, progressive deterioration of mental functions including learning and memory, and behavioral changes [1,2]. More than 35 million people across the globe today are affected by dementia, most suffering from AD, and this numbers are expected to 66 million by 2030 [3]. Although much progress has been made in understanding the cause of AD, unfortunately, there is no definitive solution for curing this disorder [4,5]. Currently, the therapeutic approaches conventionally used to maintain cognitive functions of patients consists of two classes of medications, the acetylcholinesterase inhibitors (AChEIs; tacrine, donepezil, rivastigmine, galantamine) and the noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonist (memantine), which have been approved by the US FDA for the treatment of AD [6,7]. However, these drugs exhibited mild ameliorating effect on cognitive symptoms, limited duration of their efficacy and some undesired effects, such as diarrhea, vomiting, nausea, and weight loss for AChEIs, and constipation, confusion, dizziness, and headache for memantine, respectively [8]. Due to the complexity of pathology of AD and the limitations of current AD therapeutics, increasing attentions on the development of innovative therapeutics have become urgently. The neuropathological features of AD include the excessive extracellular deposition of amyloid-β (Aβ) protein in senile plaques, hyperphosphorylated tau-containing neurofibrillary tangle, and massive neuronal death [9]. Aβ is a 39–43 amino acid peptide fragment derived from amyloid precursor protein by proteolytic processing with β-secretase and γ-secretase [10]. Experimental data from in vitro and in vivo studies indicated that Aβ can elicit apoptosis in neurons, suggesting a critical role of Aβ in the pathogenesis of AD, though the precise mechanism underlying this Aβ neurotoxicity remains to be fully elucidated [11]. In this regard, therapeutic strategies that address the modulation of Aβ toxicity may foster novel developments for the treatment of AD. Thus, searching for any substances from natural sources that can block Aβ toxicity on a cultured neuronal cell line may provide clinical benefits to control the onset of AD patients.

There is a long history of the use of traditional Chinese medicinal medicine (TCM) in the treatment of neurological disorders and these effects were documented in Chinese herbal books [12], suggesting that TCM may be a valuable resource of alternative and complementary candidates for the treatment of AD. Until now, TCM is still in current use and has been attracted a great deal of attention in recent years. For example, Chinese herbal extracts have been found to be effective in the treatment of age-associated neurodegenerative diseases, including Angelica sinensis [13], Polygonum multiflorum [14], Gastrodia elata [3], Eriobotrya japonica [15], and so on. Corydalis yanhusuo W.T. Wang is a well-known TCM that has been used traditionally to promote blood circulation, reinforce vital energy and relieve pain for a long history [16,17]. Alkaloids are known to be its main pharmacologically active ingredients in C. yanhusuo [18], and modern pharmacological experiments have recently shown that these have extensive biological functions such as antitumor, antinociceptive, antihypertensive, and antimyocardial ischemia [19]. Recently, several lines of evidence demonstrated that C. yanhusuo possessed certain beneficial effect on neurodegenerative patients with cognitive impairment, cerebral circulation insufficiency or dementia [20,21,22]. Up to now, most research on C. yanhusuo has been extensively focused on alkaloid and its derivatives, but few reports has been initiated on the polysaccharide from this plant, except that a water soluble polysaccharide has been isolate and purified from it. Considering its potential benefit on the neurodegenerative disease and no available information about the protective effect of its polysaccharide constituents on nervous system, especially at the cellular level, in this study, we aim to purify another important water-soluble neutral polysaccharide from this plant and observe its effect against Aβ-induced toxicity in PC 12 cells. Furthermore, the underlying molecular signaling pathway involved in the neuroprotective effect of this polysaccharide on Aβ-induced neurotoxicity in PC12 cells was also elucidated.