- Research article
- Open Access
- Open Peer Review
This article has Open Peer Review reports available.
High molecular weight of polysaccharides from Hericium erinaceus against amyloid beta-induced neurotoxicity
© The Author(s). 2016
Received: 13 October 2015
Accepted: 28 May 2016
Published: 7 June 2016
Hericium erinaceus (HE) is a well-known mushroom in traditional Chinese food and medicine. HE extracts from the fruiting body and mycelia not only exhibit immunomodulatory, antimutagenic and antitumor activity but also have neuroprotective properties. Here, we purified HE polysaccharides (HEPS), composed of two high molecular weight polysaccharides (1.7 × 105 Da and 1.1 × 105 Da), and evaluated their protective effects on amyloid beta (Aβ)-induced neurotoxicity in rat pheochromocytoma PC12 cells.
HEPS were prepared and purified using a 95 % ethanol extraction method. The components of HEPS were analyzed and the molecular weights of the polysaccharides were determined using high-pressure liquid chromatography (HPLC). The neuroprotective effects of the polysaccharides were evaluated through a 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay and an MTT assay and by quantifying reactive oxygen species (ROS) and mitochondrial membrane potentials (MMP) of Aβ-induced neurotoxicity in cells.
Our results showed that 250 μg/ml HEPS was harmless and promoted cell viability with 1.2 μM Aβ treatment. We observed that the free radical scavenging rate exceeded 90 % when the concentration of HEPS was higher than 1 mg/mL in cells. The HEPS decreased the production of ROS from 80 to 58 % in a dose-dependent manner. Cell pretreatment with 250 μg/mL HEPS significantly reduced Aβ-induced high MMPs from 74 to 51 % and 94 to 62 % at 24 and 48 h, respectively. Finally, 250 μg/mL of HEPS prevented Aβ-induced cell shrinkage and nuclear degradation of PC12 cells.
Our results demonstrate that HEPS exhibit antioxidant and neuroprotective effects on Aβ-induced neurotoxicity in neurons.
Hericium erinaceus (HE) is a well-known mushroom that is consumed as food and used in traditional Chinese medicine. These mushrooms contain physiologically significant components, such as β-glucan polysaccharides and other biomaterials, which have demonstrated anticancer, immunomodulatory, hypolipidemic, antioxidant and neuroprotective properties [1–6]. As an anticancer agent, the polysaccharides from HE have more significant anti-artificial pulmonary metastatic tumor effects and immunomodulatory activity than those of Hericium laciniatum . HE and Lentinus edodes have been compared with regard to their antitumor activities and immunoregulatory effects on mice with sarcoma 180 . Additionally, HE extracts (HTJ5 and HTJ5A) have been found to be more effective and less toxic than clinically used anticancer drugs such as 5-fluorouracil against liver cancer HepG2 and Huh-7, colon cancer HT-29 and gastric cancer NCI-87 cells in vitro and in tumor xenografts in vivo .
Macrophages are activated by HE polysaccharides to produce nitric oxide and express cytokines (IL-1beta and TNF-beta), which lead to effective antitumor activity and immunomodulation . Previously, we demonstrated that HE extracts can induce the activation of dendritic cells and increase the secretion of IL-12 to modulate a TH1 immune response . The hypolipidemic effects proportionally increased with oral administration of an HE exo-biopolymer in a dose-dependent manner in animal studies . The HE biomaterials reduced levels of low-density lipoprotein cholesterol while maintaining relatively high levels of high-density lipoprotein cholesterol and reduced the risk of atherosclerosis.
It was previously reported that HE extracts have neuroprotective effects, promote normal development of cultivated cerebellar cells and have regulatory effects on the development of myelin genesis processes in vitro . The ethanol extract of HE has been shown to induce nerve growth factor expression and to prevent Aβ25–35-induced impairment of memory functions in animal experiments [11, 12]. Oxidative stress has been shown to be involved in the initiation and progression of various disorders caused by oxygen radicals, which damages lipids, proteins and nucleic acids [13, 14]. The hot water extract of HE has been reported to improve this free radical scavenging activity and inhibit lipid peroxidation . HE polysaccharide extracts have been reported to decrease lipid peroxidation levels, increase antioxidant enzyme activity and increase radical scavenging activity [4, 16, 17].
In this study, we purified HEPS, which consists of two high molecular weight polysaccharides and exhibits antioxidant activity, from fruiting bodies. HEPS-treated cells showed an increase in the rate of free radical scavenging, a reduction in the production of ROS, a recovery in mitochondrial function, maintenance in morphology changes, and a reduction in cell apoptosis of PC12 cells upon Aβ treatment. Finally, we demonstrated that HEPS has neuroprotective properties for neurons.
PC12 cells were purchased from the Bioresource Collection and Research Center of the Food Industry and Development Research Institute in Taiwan. Cells were grown in RPMI 1640 with 10 % heat inactivated horse serum, 5 % fetal bovine serum, penicillin (50 units/ml), and streptomycin (50 mg/ml). The cells were cultivated in an incubator with 5 % CO2 at 37 °C.
Preparation of HEPS
Fresh fruiting bodies of HE were obtained from a local farm as previously reported . Samples of HE were identified by Professor Wen-Te Chang of China Medical University (CMU) in Taiwan. The HE voucher specimen and number (CPSCMU HE 1021202) were deposited to the School of Chinese Medicine Resources (SCMR) at CMU. A modified procedure from Dr. Mori’s report was used to prepare HEPS . The whole fruiting body was cleaned, lyophilized and powdered. The HE powder was mixed with two volumes of ethanol (95 %) and homogenized at 200 rpm for 1 h. This procedure was repeated three times. The mixture was then filtered with Whatman filter paper (Sigma-Aldrich, USA), and the extract was collected by centrifuging the mixture at 10,000 × g for 10 min at 4 °C. The HEPS supernatant was then lyophilized and stored at −20 °C until used for experiments.
Measuring components and molecular weights of HEPS
The total sugars and reducing sugars in the extract were measured as previously described . The Bradford method was used to determine the total concentration of protein using a protein assay kit (Bio-Rad, USA) following the manufacturer’s instructions. Flavonoids from HEPS were measured using previously described methods . Flavonoid content curves were determined using quercetin as a standard. The endotoxicity of HEPS was measured using a chromogenic Limulus amebocyte lysate kit (Associates of Cape Cod, USA), where the maximum sensitivity level was 0.25 EU/mL .
The molecular weights of HEPS components were determined by HPLC analysis. The extract was dissolved in deionized water, filtered through a 0.45 μm membrane and applied to a Hitachi L-2490 HPLC system (Tokyo, Japan) as a 20 μL aliquot. The system was fit with a TSK-GEL G3000PWXL column (7.8 mm × 30 cm) and was maintained at a temperature of 25 °C. The extract was eluted with deionized water at a flow rate of 0.6 mL/min and detected by a refractive index detector (RID). Pullulan standards of various molecular weights (5900, 11,800, 22,800, 47,300, and 112,000 daltons) were used to establish standard curves and to determine molecular weights .
DPPH radical scavenging assay
The free radical scavenging rate was evaluated by measuring the 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity of HEPS. The DPPH assay used a modified procedure from a previously described study . The HE extracts were dissolved in methanol and mixed with 250 μL of a 0.2 mM DPPH radical solution (Sigma-Aldrich, USA). After 30 min at room temperature, the absorbance of the resulting solutions and a blank were recorded against 0.1 mg/mL butylated hydroxyanisole (BHA) and L-ascorbic acid (Vitamin C; Sigma-Aldrich, USA) as positive controls. The absorbance of each reaction was recorded in triplicate. The disappearance of DPPH radicals was measured spectrophotometrically at 517 nm using a Hitachi U-2001 spectrophotometer (Tokyo, Japan), and the DPPH scavenging effect was calculated as previously described .
MTT assay for cell cytotoxicity and protection
The MTT assay was used for three experiments. First, the cell cytotoxicity of HEPS was measured by plating exponentially growing PC12 cells at a density of 5 × 104 cells/well in 96-well plates, which were exposed with or without 25, 50, 100, 200, 250 μg/mL of HEPS for 24 and 48 h. The second stage of the assay measured cell cytotoxicity of Aβ1 - 40 (Sigma-Aldrich, USA) by adding 1.2 μM Aβ1 - 40 to PC12 cells for 24 and 48 h. The third stage was a cell protection assay, in which PC12 cells were incubated with 25, 50, 100, 200, 250 μg/mL of HEPS for 24 h, and 1.2 μM Aβ1 - 40 was added for 24 and 48 h. After each of these three experiments, the cells were incubated with 2 mg/mL 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) for 4 h at 37 °C, the media was carefully removed and 100 μl of DMSO was added to each well. Dark blue formazan crystals formed, the intact cells were solubilized for 30 min, and the absorbance at 570 nm was measured with a PowerWave XS ELISA reader (Bio-Tek, USA). The results were expressed as the percentage of MTT reduction, assuming the absorbance of control cells was 100 %.
ROS and MMP measurements
To measure ROS, cells treated with HEPS and Aβ1 - 40 were collected and centrifuged at 650 × g for 10 min. The resulting pellets were washed once with phosphate buffered saline (PBS). These steps were repeated twice. The ROS production rate was measured using an OxiSelect™ Intracellular ROS Assay Kit, and the intracellular accumulation of ROS was monitored using the cell-permeable fluorogenic probe 2’,7’-dichlorodihydrofluorescein diacetate (DCFH-DA).
The MMP was measured using the fluorescent dye JC-1 . Mitochondria with high MMP promoted the formation of J-aggregates and fluoresced red. In contrast, mitochondria with low MMP contained JC-1 monomers and fluoresced green. After co-treating cells with 1.2 μM Aβ1 - 40 for 24 h in the presence or absence of HEPS, 1 × 106 cells/mL were collected and incubated for 15 min at 37 °C. JC-1 (10 μg/mL) was then loaded, and the fluorescence intensity of the cells was examined at an excitation of 485 nm and emission of 535 nm using FACScan flow cytometry (Becton Dickinson, USA).
Cell morphology and intracellular fluorescence staining
The DNA-binding dye acridine orange (Sigma-Aldrich, USA) was used to observe the morphological characteristics of the treated cells. After PC12 cells were incubated with 1.2 μM Aβ1 - 40 or HEPS at 37 °C for 24 and 48 h, the cells were washed with sterilized PBS three times and incubated with acridine orange (10 μg/ml) at 37 °C for 10 min in the dark. The stained cells were observed and photographed using an Olympus COVER-018 fluorescence microscope (Tokyo, Japan).
The data were analyzed using Statistical Analysis System (SAS) software (SAS Institute, USA) as described previously . A one-way analysis of variance (one-way ANOVA) and Duncan’s test were used to determine the statistical significance between groups. Differences were considered statistically significant when p ≤ 0.05.
Results and discussion
The composition and cell toxicity of HEPS
Compositions of the extract from Hericium erinaceus
311 ± 27.8a
249 ± 25.8a
135 ± 0.1a
99 ± 1.7a
HEPS protecting PC12 cells against Aβ1–40 induced neurotoxicity
HEPS inhibited accumulation of free radical and ROS in cells
Mitochondria are a major source of ROS, which are produced in many normal and abnormal physiological processes . However, excessive ROS production may cause damage during the accumulation of Aβ in the pathogenesis of AD . As shown in Fig. 4b, pretreatment of HEPS at concentrations ranging from 25 μg/mL to 250 μg/mL significantly decreased the production of ROS from 80 to 58 % after Aβ incubation for 24 h. Moreover, 250 μg/mL HEPS considerably reduced ROS levels to 40 % compared to cells without HEPS pretreatment, suggesting that HEPS protects mitochondria and reduces ROS generation.
HEPS prevents loss of MMP in PC12 cells
Measurement of morphology and intracellular changes
The compound CBNU06 is purified from Isodon japonicas and protects PC12 cells from Aβ-induced neurotoxicity and reduces the number of cells that undergo DNA condensation and fragmentation by inhibiting NF-kB signaling pathways . Atractylodes macrocephala polysaccharides have demonstrated neuroprotective effects by decreasing the expression of Bax and Caspase-3 and increasing Bcl-2 levels in neurons . However, the actual mechanism of protecting and reducing cell apoptosis by HEPS needs further investigation.
Our results demonstrate that pretreatment of PC12 cells with HEPS, which contains two high molecular weight polysaccharides, promotes antioxidant activity and has neuroprotective effects against on Aβ-induced neurotoxicity. We show that HEPS promoted cell viability under Aβ-induced toxic conditions. Furthermore, HEPS also increased the efficacy of free radical scavenging and ROS. Finally, HEPS protected PC12 cells against Aβ-induced cell apoptosis. In summary, our previous and current findings suggest that different molecular weight polysaccharides from HE not only play a role in immunomodulation of dendritic cells but also contain neuroprotective effects for neurons.
AD, Alzheimer's disease; APP, amyloid precursor protein; Aβ, amyloid beta; BHA, butylated hydroxyanisole; DCFH-DA, 2’, 7’-Dichlorodihydrofluorescin diacetate; DPPH, 2,2-diphenyl-1-picrylhydrazil; GSH, glutathione; GSSG, oxidized glutathione; GST, glutathione S-transferase; HE, Hericium erinaceus; HPLC, high pressure liquid chromatography; MMP, mitochondrial membrane potential; MTT, 3- (4,5-cimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide; PBS, phosphate buffered saline; RID, refractive index detector; ROS, reactive oxygen species; SAS, Statistical Analysis System
The authors would like to thank all of the colleagues and students who contributed to this study.
This study is partially supported by a grant from the Chang Gung Research Foundation (CMRPG 8C1351).
Availability of data and materials
The datasets supporting the conclusions of this article are included within this article.
JHC, SCS, and MSL participated in this study with primary duties including the conception and design of the study, data analysis, data interpretation, drafting the article and final approval of the version to be submitted. CLT participated in this study with primary duties in reference searches, data analysis and data interpretation. YYL participated in this study with primary duties in data acquisition, data analysis, reference searches and final approval of the version to be submitted.
The authors declare that they have no competing interests.
Consent for publication
Ethics approval and consent to participate
Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
- Wang JC, Hu SH, Su CH, Lee TM. Antitumor and immunoenhancing activities of polysaccharide from culture broth of Hericium spp. Kaohsiung J Med Sci. 2001;17(9):461–7.PubMedGoogle Scholar
- Sheu S-C, Lyu Y, Lee M-S, Cheng J-H. Immunomodulatory effects of polysaccharides isolated from Hericium erinaceus on dendritic cells. Process Biochem. 2013;48(9):1402–8.View ArticleGoogle Scholar
- Yang BK, Park JB, Song CH. Hypolipidemic effect of an Exo-biopolymer produced from a submerged mycelial culture of Hericium erinaceus. Biosci Biotechnol Biochem. 2003;67(6):1292–8.View ArticlePubMedGoogle Scholar
- Wong KH, Sabaratnam V, Abdullah N, Kuppusamy UR, Naidu M. Effects of cultivation techniques and processing on antimicrobial and antioxidant activities of hericium erinaceus (Bull.:Fr.) Pers. Extracts. Food Technol Biotechnol. 2009;47(1):47–55.Google Scholar
- Lai PL, Naidu M, Sabaratnam V, Wong KH, David RP, Kuppusamy UR, et al. Neurotrophic properties of the Lion's mane medicinal mushroom, Hericium erinaceus (Higher Basidiomycetes) from Malaysia. Int J Med Mushrooms. 2013;15(6):539–54.View ArticlePubMedGoogle Scholar
- Khan MA, Tania M, Liu R, Rahman MM. Hericium erinaceus: an edible mushroom with medicinal values. J Complement Integr Med. 2013;10. doi:10.1515/jcim-2013-0001.
- Liu C, Gao P, Qian J, Yan W. Immunological study on the antitumor effects of fungus polysaccharides compounds. Wei Sheng Yan Jiu. 2000;29(3):178–80.PubMedGoogle Scholar
- Li G, Yu K, Li F, Xu K, Li J, He S, et al. Anticancer potential of Hericium erinaceus extracts against human gastrointestinal cancers. J Ethnopharmacol. 2014;153(2):521–30.View ArticlePubMedGoogle Scholar
- Lee JS, Min KM, Cho JY, Hong EK. Study of macrophage activation and structural characteristics of purified polysaccharides from the fruiting body of Hericium erinaceus. J Microbiol Biotechnol. 2009;19(9):951–9.View ArticlePubMedGoogle Scholar
- Kolotushkina EV, Moldavan MG, Voronin KY, Skibo GG. The influence of Hericium erinaceus extract on myelination process in vitro. Fiziol Zh. 2003;49(1):38–45.PubMedGoogle Scholar
- Mori K, Obara Y, Hirota M, Azumi Y, Kinugasa S, Inatomi S, et al. Nerve growth factor-inducing activity of Hericium erinaceus in 1321N1 human astrocytoma cells. Biol Pharm Bull. 2008;31(9):1727–32.View ArticlePubMedGoogle Scholar
- Mori K, Obara Y, Moriya T, Inatomi S, Nakahata N. Effects of Hericium erinaceus on amyloid beta(25–35) peptide-induced learning and memory deficits in mice. Biomed Res. 2011;32(1):67–72.View ArticlePubMedGoogle Scholar
- Aruoma OI, Kaur H, Halliwell B. Oxygen free radicals and human diseases. J R Soc Health. 1991;111(5):172–7.View ArticlePubMedGoogle Scholar
- Ames BN, Shigenaga MK, Hagen TM. Oxidants, antioxidants, and the degenerative diseases of aging. Proc Natl Acad Sci U S A. 1993;90(17):7915–22.View ArticlePubMedPubMed CentralGoogle Scholar
- Abdullah N, Ismail SM, Aminudin N, Shuib AS, Lau BF. Evaluation of selected culinary-medicinal mushrooms for antioxidant and ACE inhibitory activities. Evid Based Complement Alternat Med. 2012;2012:464238.View ArticlePubMedGoogle Scholar
- Zhang Z, Lv G, Pan H, Pandey A, He W, Fan L. Antioxidant and hepatoprotective potential of endo-polysaccharides from Hericium erinaceus grown on tofu whey. Int J Biol Macromol. 2012;51(5):1140–6.View ArticlePubMedGoogle Scholar
- Han ZH, Ye JM, Wang GF. Evaluation of in vivo antioxidant activity of Hericium erinaceus polysaccharides. Int J Biol Macromol. 2013;52:66–71.View ArticlePubMedGoogle Scholar
- Zhishen J, Mengcheng T, Jianming W. The determination of flavonoid contents in mulberry and their scavenging effects on superoxide radicals. Food Chem. 1999;64(4):555–9.View ArticleGoogle Scholar
- Shimada K, Fujikawa K, Yahara K, Nakamura T. Antioxidative properties of xanthan on the autoxidation of soybean oil in cyclodextrin emulsion. J Agric Food Chem. 1992;40(6):945–8.View ArticleGoogle Scholar
- Reers M, Smiley ST, Mottola-Hartshorn C, Chen A, Lin M, Chen LB. Mitochondrial membrane potential monitored by JC-1 dye. Methods Enzymol. 1995;260:406–17.View ArticlePubMedGoogle Scholar
- Yao LH, Jiang YM, Shi J, Tomas-Barberan FA, Datta N, Singanusong R, et al. Flavonoids in food and their health benefits. Plant Foods Hum Nutr. 2004;59(3):113–22.View ArticlePubMedGoogle Scholar
- Vauzour D, Vafeiadou K, Rodriguez-Mateos A, Rendeiro C, Spencer JP. The neuroprotective potential of flavonoids: a multiplicity of effects. Genes Nutr. 2008;3(3–4):115–26.View ArticlePubMedPubMed CentralGoogle Scholar
- Vauzour D. Effect of flavonoids on learning, memory and neurocognitive performance: relevance and potential implications for Alzheimer's disease pathophysiology. J Sci Food Agric. 2014;94(6):1042–56.View ArticlePubMedGoogle Scholar
- Giavasis I. Bioactive fungal polysaccharides as potential functional ingredients in food and nutraceuticals. Curr Opin Biotechnol. 2014;26:162–73.View ArticlePubMedGoogle Scholar
- Ho YS, Yu MS, Yang XF, So KF, Yuen WH, Chang RC. Neuroprotective effects of polysaccharides from wolfberry, the fruits of Lycium barbarum, against homocysteine-induced toxicity in rat cortical neurons. J Alzheimers Dis. 2010;19(3):813–27.PubMedGoogle Scholar
- Chen G, Chen KS, Knox J, Inglis J, Bernard A, Martin SJ, et al. A learning deficit related to age and beta-amyloid plaques in a mouse model of Alzheimer's disease. Nature. 2000;408(6815):975–9.View ArticlePubMedGoogle Scholar
- Walsh DM, Klyubin I, Fadeeva JV, Cullen WK, Anwyl R, Wolfe MS, et al. Naturally secreted oligomers of amyloid beta protein potently inhibit hippocampal long-term potentiation in vivo. Nature. 2002;416(6880):535–9.View ArticlePubMedGoogle Scholar
- Zhou ZY, Tang YP, Xiang J, Wua P, Jin HM, Wang Z, et al. Neuroprotective effects of water-soluble Ganoderma lucidum polysaccharides on cerebral ischemic injury in rats. J Ethnopharmacol. 2010;131(1):154–64.View ArticlePubMedGoogle Scholar
- Liu Y, Fukuwatari Y, Okumura K, Takeda K, Ishibashi K, Furukawa M, et al. Immunomodulating activity of Agaricus brasiliensis KA21 in mice and in human volunteers. Evid Based Complement Alternat Med. 2008;5(2):205–19.View ArticlePubMedGoogle Scholar
- Park SY. Potential therapeutic agents against Alzheimer's disease from natural sources. Arch Pharm Res. 2010;33(10):1589–609.View ArticlePubMedGoogle Scholar
- Yu MS, Leung SK, Lai SW, Che CM, Zee SY, So KF, et al. Neuroprotective effects of anti-aging oriental medicine Lycium barbarum against beta-amyloid peptide neurotoxicity. Exp Gerontol. 2005;40(8–9):716–27.View ArticlePubMedGoogle Scholar
- Chen W, Cheng X, Chen J, Yi X, Nie D, Sun X, et al. Lycium barbarum polysaccharides prevent memory and neurogenesis impairments in scopolamine-treated rats. PLoS One. 2014;9(2):e88076.View ArticlePubMedPubMed CentralGoogle Scholar
- Tsai M-C, Song T-Y, Shih P-H, Yen G-C. Antioxidant properties of water-soluble polysaccharides from Antrodia cinnamomea in submerged culture. Food Chem. 2007;104(3):1115–22.View ArticleGoogle Scholar
- Malinowska E, Krzyczkowski W, Herold F, Łapienis G, Ślusarczyk J, Suchocki P, et al. Biosynthesis of selenium-containing polysaccharides with antioxidant activity in liquid culture of Hericium erinaceum. Enzyme Microb Technol. 2009;44(5):334–43.View ArticleGoogle Scholar
- Valko M, Leibfritz D, Moncol J, Cronin MT, Mazur M, Telser J. Free radicals and antioxidants in normal physiological functions and human disease. Int J Biochem Cell Biol. 2007;39(1):44–84.View ArticlePubMedGoogle Scholar
- Mohsenzadegan M, Mirshafiey A. The immunopathogenic role of reactive oxygen species in Alzheimer disease. Iran J Allergy Asthma Immunol. 2012;11(3):203–16.PubMedGoogle Scholar
- Wang X, Su B, Siedlak SL, Moreira PI, Fujioka H, Wang Y, et al. Amyloid-beta overproduction causes abnormal mitochondrial dynamics via differential modulation of mitochondrial fission/fusion proteins. Proc Natl Acad Sci U S A. 2008;105(49):19318–23.View ArticlePubMedPubMed CentralGoogle Scholar
- Abramov AY, Canevari L, Duchen MR. Beta-amyloid peptides induce mitochondrial dysfunction and oxidative stress in astrocytes and death of neurons through activation of NADPH oxidase. J Neurosci. 2004;24(2):565–75.View ArticlePubMedGoogle Scholar
- Reddy PH. Amyloid beta, mitochondrial structural and functional dynamics in Alzheimer's disease. Exp Neurol. 2009;218(2):286–92.View ArticlePubMedPubMed CentralGoogle Scholar
- Eckert A, Keil U, Scherping I, Hauptmann S, Muller WE. Stabilization of mitochondrial membrane potential and improvement of neuronal energy metabolism by Ginkgo biloba extract EGb 761. Ann N Y Acad Sci. 2005;1056:474–85.View ArticlePubMedGoogle Scholar
- Roth AD, Ramirez G, Alarcon R, Von Bernhardi R. Oligodendrocytes damage in Alzheimer's disease: beta amyloid toxicity and inflammation. Biol Res. 2005;38(4):381–7.View ArticlePubMedGoogle Scholar
- Kim HS, Lim JY, Sul D, Hwang BY, Won TJ, Hwang KW, et al. Neuroprotective effects of the new diterpene, CBNU06 against beta-amyloid-induced toxicity through the inhibition of NF-kappaB signaling pathway in PC12 cells. Eur J Pharmacol. 2009;622(1–3):25–31.View ArticlePubMedGoogle Scholar
- Hu WX, Xiang Q, Wen Z, He D, Wu XM, Hu GZ. Neuroprotective effect of Atractylodes macrocephalaon polysaccharides in vitro on neuronal apoptosis induced by hypoxia. Mol Med Rep. 2014;9(6):2573–81.PubMedGoogle Scholar